Human Phospho-FLT3 (Y589) and Total FLT3 ELISA

RayBio® Human Phospho-FLT3 (Tyr589) and Total FLT3 ELISA Kit. This assay semi-quantitatively measures FLT3 phosphorylated at Tyrosine-589 as well as total FLT3 in cell lysate samples.

Specifications

Product Features

• - Rapidly measure phosphorylated protein in lysates

• - Screen numerous different cell lysates without performing a Western Blot analysis

• - Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - HRP-Conjugated Secondary Antibody

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

Jurkat cells were treated with Pervanadate. Solubilize cells at 4 x 10^7 cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

Pervanadate Stimulation of A431 Cell Line

A431 cells were treated or untreated with Pervanadate. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment.

Human/Mouse/Rat Phospho-GSK3b (S9) ELISA

RayBio® Human Phospho-GSK3b (S9) ELISA Kit. This assay semi-quantitatively measures phosphorylated GSK3b (Ser9) in lysate samples.

Specifications

Introduction

RayBio? Phospho-GSK3beta (S9) ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in human, mouse and rat cell lysates. By determining phosphorylated GSK3beta protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blotting analysis.

This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human/mouse/rat phosphoGSK3beta. An anti-pan GSK3beta antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and GSK3beta present in a sample is bound to the wells by the immobilized antibody. The wells are washed and rabbit anti-GSK3beta (S9) antibody is used to detect phosphorylated GSK3beta. After washing away unbound antibody, HRP-conjugated anti-rabbit IgG is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of GSK3beta (S9). The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Product Features

• Rapidly measure phosphorylated protein in lysates
• Screen numerous different cell lysates without performing a Western Blot analysis
• Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate
• - Wash Buffer
• - Anti-Phospho Antibody
• - HRP-Conjugated Secondary Antibody
• - Assay Diluent
• - TMB One-Step Substrate
• - Stop Solution
• - Lysis Buffer
• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water
• - 100 ml and 1 liter graduated cylinders
• - Tubes to prepare sample dilutions
• - Protease and Phosphatase inhibitors
• - Precision pipettes to deliver 2 µl to 1 ml volumes
• - Adjustable 1-25 ml pipettes for reagent preparation
• - Benchtop rocker or shaker
• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.
• 2. Add 100 µl of sample or positive control to each well.
• 3. Incubate 2.5 h at RT or O/N at 4 °C.
• 4. Add 100 µl of prepared primary antibody to each well.
• 5. Incubate 1 h at RT.
• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.
• 7. Incubate 1 h at RT.
• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.
• 9. Incubate 30 min at RT.
• 10. Add 50 µl of Stop Solution to each well.
• 11. Read at 450 nm immediately.

Typical Data

Positive Control

NIH 3T3 cells were treated with recombinant human PDGF BB at 37oC for 10 min. Solubilize cells at 4 x 10⁷ cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

Recombinant Human PDGF BB Stimulation of NIH 3T3 Cell Lines

NIH 3T3 cells were treated or untreated with 50 ng/ml recombinant human PDGF BB for 10 min. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4°C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge, and store at –20°C. Item D, store at 2-8°C for up to one month (store at -20°C for up to 6 months, avoid repeated freeze-thaw cycles). Reconstituted Positive Control (Item K) should be stored at -70°C.

Human Phospho-eIF-4E (S209) and Total eIF-4E ELISA

RayBio® Human Phospho-eIF-4E (S209) and Total eIF-4E ELISA Kit. This assay semi-quantitatively measures phosphorylated eIF-4E (Ser209) and Total eIF-4E in lysate samples.

Specifications

Introduction

RayBio® Phospho-eIF4E (Ser209) and Total eIF4E ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in human cell lysates. By determining phosphorylated eIF4E protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blotting analysis.

This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human phospho-eIF4E and total eIF4E. An anti-pan eIF4E antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and eIF4E present in a sample is bound to the wells by the immobilized antibody. The wells are washed and rabbit anti-eIF4E (Ser209) antibody is used to detect phosphorylated eIF4E or rabbit anti-eIF4E antibody is used to detect pan eIF4E. After washing away unbound antibody, HRP-conjugated anti-rabbit IgG is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of eIF4E (Ser209) or pan eIF4E bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Product Features

• - Rapidly measure phosphorylated protein in lysates

• - Screen numerous different cell lysates without performing a Western Blot analysis

• - Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - Anti-Pan Antibody

• - HRP-Conjugated Secondary Antibody

• - Streptavidin-Conjugated HRP

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

HeLa cells were treated with Anisomycin. Solubilize cells at 4 x 10⁷ cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

Anisomycin Stimulation of HeLa Cell Lines

HeLa cells were treated or untreated with Anisomycin. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4°C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge, and store at –20°C. Item D, store at 2-8°C for up to one month (store at -20°C for up to 6 months, avoid repeated freeze-thaw cycles). Reconstituted Positive Control (Item K) should be stored at -70°C.

Human Phospho-FYN (Y530) and Total FYN ELISA

RayBio® Human Phospho-FYN (Tyr530) and Total FYN ELISA Kit. This assay semi-quantitatively measures FYN phosphorylated at Tyrosine-530 as well as total FYN in lysate samples.

Specifications

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - HRP-Conjugated Secondary Antibody

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

Jurkat cells were treated with Pervanadate. Solubilize cells at 4 x 10⁷ cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

Pervanadate (PV) Stimulation of 293 Cell Line

293 cells were treated or untreated with Pervanadate. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment.

Human Phosphotyrosine EphA4 ELISA

RayBio® Human Phosphotyrosine EphA4 ELISA Kit. This assay semi-quantitatively measures phosphotyrosine EphA4 in lysate samples.

Specifications

Introduction

RayBio^® Phospho-EphA4 ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in cell lysates. By determining phospho-EphA4 in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blot analysis.

This sandwich-based ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human phospho-EphA4. An anti-EphA4 antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and phosphorylated and unphosphorylated EphA4 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-phosphotyrosine antibody is used to detect only tyrosine-phosphorylated protein. After washing away unbound antibody, HRP-conjugated streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of phosphor-EphA4 bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Product Features

• - Rapidly measure phosphorylated protein in lysates

• - Screen numerous different cell lysates without performing a Western Blot analysis

• - Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Biotinylated Anti-Phosphotyrosine Antibody

• - Stop Solution

• - Assay Diluent(s)

• - Positive Control Sample

• - Lysis Buffer

• - Streptavidin-Conjugated HRP

• - TMB One-Step Substrate

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

Jurkat cells were treated with Pervanadate at 37°C for 10 min. Solubilize cells at 4 x 10⁷ cells/ml in lysis buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for details.

Pervanadate Stimulation of Jurkat Cell Line

Jurkat cells were treated or untreated with Pervanadate for 10 min at 37°C. Cell lysates were analyzed using this phosphoELISA.

Storage/Stability

Upon receipt, the kit should be stored at -20 °C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), HRP-Streptavidin (Item G), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4 °C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge and store at -20 °C. Reconstituted Positive Control (Item K) should be stored at -70 °C.

Human/Mouse Phospho-p38 alpha (T180/Y182) ELISA

RayBio® Human/Mouse Phospho-p38 alpha (T180/Y182) ELISA Kit. This assay semi-quantitatively measures phosphorylated p38 alpha (Thr180/Tyr182) in lysate samples.

Specifications

Product Features

• - Rapidly measure phosphorylated protein in lysates

• - Screen numerous different cell lysates without performing a Western Blot analysis

• - Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - HRP-Conjugated Secondary Antibody

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Storage/Stability

Upon receipt, the kit should be stored at -20 °C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), HRP-Streptavidin (Item G), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4 °C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge and store at -20 °C. Reconstituted Positive Control (Item K) should be stored at -70 °C.

Citations

Bizargity P, Liu K, Wang L, Hancock WW, Visner GA. Inhibitory Effects of Pirfenidone on Dendritic Cells and Lung Allograft Rejection. Transplantation. 2012;94(2):114-122. doi:10.1097/TP.0b013e3182584879.

Species: Mouse

Sample Type: Cell Lysate (Lipopolysaccharide and allogeneic treated mice bone marrow DC)

Human/Mouse/Rat Phospho-SHP2 (Y542) ELISA

RayBio® Human/Mouse/Rat Phospho-SHP2 (Y542) ELISA Kit. This assay semi-quantitatively measures phosphorylated SHP2 (Tyr542) in lysate samples.

Specifications

Introduction

RayBio® Phospho- SHP2 (Y542) ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in human, mouse and rat cell lysates. By determining phosphorylated SHP2 protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blotting analysis.

This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human, mouse and rat phospho-SHP2. An anti-pan SHP2 antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and SHP2 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and rabbit anti-SHP2 (Y542) antibody is used to detect phosphorylated SHP2. After washing away unbound antibody, HRP-conjugated anti-rabbit IgG is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of SHP2 (Y542) bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Product Features

• Rapidly measure phosphorylated protein in lysates

• Screen numerous different cell lysates without performing a Western Blot analysis

• Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - HRP-Conjugated Secondary Antibody

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• -  Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

NIH3T3 cells were treated with PDGFBB at 37°C for 10 min. Solubilize cells at 4 x 10⁷ cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

PDGFBB Stimulation of NIH3T3 Cell Line

NIH3T3 cells were treated or untreated with 50ng/ml PDGFBB for 10 min. Cell lysates were analyzed using this phospho ELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4°C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge, and store at –20°C. Item D, store at 2-8°C for up to one month (store at -20°C for up to 6 months, avoid repeated freeze-thaw cycles). Reconstituted Positive Control (Item K) should be stored at -70°C.

Human/Mouse/Rat Phospho-P53 (S15) ELISA

Phospho-P53 (S15) ELISA Kit. This assay semi-quantitatively measures phosphorylated P53 (Ser15) in lysate samples.

Specifications

Introduction

RayBio® Phospho-P53 (S15) ELISA kit is a very rapid, convenient and sensitive assay kit that can monitor the activation or function of important biological pathways in human, mouse and rat cell lysates. By determining phosphorylated P53 protein in your experimental model system, you can verify pathway activation in your cell lysates. You can simultaneously measure numerous different cell lysates without spending excess time and effort in performing a Western Blotting analysis.

This Sandwich ELISA kit is an in vitro enzyme-linked immunosorbent assay for the measurement of human, mouse and rat phospho-P53. An anti-P53 (S15) antibody has been coated onto a 96-well plate. Samples are pipetted into the wells and phosphorylated P53 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-P53 antibody is used to detect phosphorylated P53. After washing away unbound antibody, HRP-conjugated streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of P53 (S15). The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm.

Product Features

• Rapidly measure phosphorylated protein in lysates

• Screen numerous different cell lysates without performing a Western Blot analysis

• Minimal hands-on time, convenient, and non-radioactive material

Application Notes

Kit Components

• - Pre-Coated 96-well Strip Microplate

• - Wash Buffer

• - Anti-Phospho Antibody

• - HRP-Conjugated Secondary Antibody

• - Assay Diluent

• - TMB One-Step Substrate

• - Stop Solution

• - Lysis Buffer

• - Positive Control Sample

Other Materials Required

• - Distilled or deionized water

• - 100 ml and 1 liter graduated cylinders

• - Tubes to prepare sample dilutions

• - Protease and Phosphatase inhibitors

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - Benchtop rocker or shaker

• - Microplate reader capable of measuring absorbance at 450 nm

Protocol Outline

• 1. Prepare all reagents and samples as instructed in the manual.

• 2. Add 100 µl of sample or positive control to each well.

• 3. Incubate 2.5 h at RT or O/N at 4 °C.

• 4. Add 100 µl of prepared primary antibody to each well.

• 5. Incubate 1 h at RT.

• 6. Add 100 µl of prepared 1X HRP-Streptavidin to each well.

• 7. Incubate 1 h at RT.

• 8. Add 100 µl of TMB One-Step Substrate Reagent to each well.

• 9. Incubate 30 min at RT.

• 10. Add 50 µl of Stop Solution to each well.

• 11. Read at 450 nm immediately.

Typical Data

Positive Control

T47D cells were exposed to 50J/m² of UV light followed by a 4 hours recovery period. Solubilize cells at 4 x 10⁷ cells/ml in Cell Lysate Buffer. Serial dilutions of lysates were analyzed in this ELISA. Please see step 3 of Part VI Reagent Preparation for detail.

UV irradiation of T47D Cell Line

T47D cells were untreated or exposed to 50J/m² of UV light followed by a 4 hours recovery period before lysis. Cell lysates were analyzed using this phosphoELISA and Western Blot.

Storage/Stability

Upon receipt, the kit should be stored at –20°C. Please use within 6 months from the date of shipment. After initial use, Wash Buffer Concentrate (Item B), Assay Diluent (Item E), TMB One-Step Substrate Reagent (Item H), Stop Solution (Item I) and Cell Lysate Buffer (Item J) should be stored at 4°C to avoid repeated freeze-thaw cycles. Return unused wells to the pouch containing desiccant pack, reseal along entire edge, and store at –20°C. Item D, store at 2-8°C for up to one month (store at -20°C for up to 6 months, avoid repeated freeze-thaw cycles). Reconstituted Positive Control (Item K) should be stored at -70°C.

Human IL-1 beta/IL-1 F2 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) IL-1 beta/IL-1 F2 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-1 Beta IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-1 Beta in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-1 Beta coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-1 Beta present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Standard Curves

Typical Data

Spiking & Recovery Results

Serum spike tests show recovery is 107% with a range of 120% to 94%

Application Notes

• - IL-1 Beta Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-1 Beta

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-1 Beta

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-1 Beta (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-1 Beta affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80^°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25µl standard or sample to each well. Incubate for 2.5 hours at room temperature or overnight at 4^°C

• 3. Add 25 µl detection affinity reagent to each well. Incubate 1 hour at room temperature

• 4. Add 25µL of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 15µL Primer solution 10µL of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2^° to 8^°C from the date of shipment. Standard (recombinant protein) should be stored at -20^°C or -80^°C (recommended at -80^°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2^° to 8^°C. Note: the kit can be used within one year if the whole kit is stored at -20^°C. Avoid repeated freeze-thaw cycles.

Human TNF alpha IQELISA™ Kit

RayBio^® Human Immunoquantitative (PCR-Based) TNF alpha ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human TNF alpha IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human TNF alpha in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human TNF alpha coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and TNF alpha present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curves

Typical Data

Spiking & Recovery Results

Serum spike tests show recovery is 125% with a range of 147% to 129%

Application Notes

Kit Components

• - TNF alpha Microplate (Item A)**: 96 well PCR plate coated with anti-Human TNF alpha

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human TNF alpha

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for TNF alpha (Item F): 2 vials of a 4x concentrated solution of anti-Human TNF alpha affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Mouse IL-10 IQELISA™ Kit

RayBio® Mouse Immunoquantitative (PCR-Based) IL-10 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Mouse IL-10 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Mouse IL-10 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Mouse IL-10 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-10 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Application Notes

• - IL-10 Microplate (Item A)**: 96 well PCR plate coated with anti-Mouse IL-10

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Mouse IL-10

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-10 (Item F): 2 vials of a 4x concentrated solution of anti-Mouse IL-10 affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-6 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) IL-6 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-6 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-6 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-6 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-6 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curves

Typical Data

Application Notes

Kit Components

• - IL-6 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-6

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-6

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-6 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-6 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human BACE-1 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) BACE-1 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human BACE-1 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human BACE-1 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human BACE-1 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and BACE-1 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - BACE-1 Microplate (Item A)**: 96 well PCR plate coated with anti-Human BACE-1

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human BACE-1

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for BACE-1 (Item F): 2 vials of a 4x concentrated solution of anti-Human BACE-1 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human proBNP IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) Pro-BNP ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human Pro-BNP IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human Pro-BNP in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human Pro-BNP coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and Pro-BNP present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - Pro-BNP Microplate (Item A)**: 96 well PCR plate coated with anti-Human Pro-BNP

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human Pro-BNP

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for Pro-BNP (Item F): 2 vials of a 4x concentrated solution of anti-Human Pro-BNP affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Mouse IL-33 IQELISA™ Kit

RayBio® Mouse Immunoquantitative (PCR-Based) IL-33 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Mouse IL-33 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Mouse IL-33 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Mouse IL-33 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-33 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - IL-33 Microplate (Item A)**: 96 well PCR plate coated with anti-Mouse IL-33

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Mouse IL-33

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-33 (Item F): 2 vials of a 4x concentrated solution of anti-Mouse IL-33 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• Prepare all reagents, samples and standards as instructed

• Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-22 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) IL-22 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-22 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-22 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-22 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-22 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - IL-22 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-22

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-22

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-22 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-22 affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human S100 A7 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) S100 A7 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human S100 A7 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human S100 A7 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human S100 A7 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and S100 A7 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - S100 A7 Microplate (Item A)**: 96 well PCR plate coated with anti-Human S100 A7

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human S100 A7

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for S100 A7 (Item F): 2 vials of a 4x concentrated solution of anti-Human S100 A7 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IgG IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) IgG ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IgG IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IgG in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IgG coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IgG present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• -  IgG Microplate (Item A)**: 96 well PCR plate coated with anti-Human IgG

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IgG

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IgG (Item F): 2 vials of a 4x concentrated solution of anti-Human IgG affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human M-CSF IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) M-CSF ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human M-CSF IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human M-CSF in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human M-CSF coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and M-CSF present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - M-CSF Microplate (Item A)**: 96 well PCR plate coated with anti-Human M-CSF

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human M-CSF

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for M-CSF (Item F): 2 vials of a 4x concentrated solution of anti-Human M-CSF affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human NRG1-alpha/HRG1-alpha IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) NRG1-alpha (HRG1-alpha) ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human NRG1-alpha (HRG1-alpha) IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human NRG1-alpha (HRG1-alpha) in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human NRG1-alpha (HRG1-alpha) coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and NRG1-alpha (HRG1-alpha) present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - NRG1-alpha (HRG1-alpha) Microplate (Item A)**: 96 well PCR plate coated with anti-Human NRG1-alpha (HRG1-alpha)

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human NRG1-alpha (HRG1-alpha)

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for NRG1-alpha (HRG1-alpha) (Item F): 2 vials of a 4x concentrated solution of anti-Human NRG1-alpha (HRG1-alpha) affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-5 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) IL-5 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-5 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-5 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-5 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-5 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curves

Typical Data

Application Notes

Kit Components

• - IL-5 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-5

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-5

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-5 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-5 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human hCG-beta IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) hCG beta ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human hCG beta IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human hCG beta in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human hCG beta coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and hCG beta present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - hCG beta Microplate (Item A)**: 96 well PCR plate coated with anti-Human hCG beta

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human hCG beta

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for hCG beta (Item F): 2 vials of a 4x concentrated solution of anti-Human hCG beta affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Mouse Fetuin A IQELISA™ Kit

RayBio® Mouse Immunoquantitative (PCR-Based) Fetuin A ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Mouse Fetuin A IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Mouse Fetuin A in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Mouse Fetuin A coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and Fetuin A present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - Fetuin A Microplate (Item A)**: 96 well PCR plate coated with anti-Mouse Fetuin A

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Mouse Fetuin A

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for Fetuin A (Item F): 2 vials of a 4x concentrated solution of anti-Mouse Fetuin A affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-10 IQELISA™ Kit

RayBio^® Human Immunoquantitative (PCR-Based) IL-10 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-10 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-10 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-10 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-10 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - IL-10 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-10

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-10

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-10 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-10 affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human Pleiotrophin/PTN IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) Pleiotrophin (PTN ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human Pleiotrophin (PTN IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human Pleiotrophin (PTN in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human Pleiotrophin (PTN coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and Pleiotrophin (PTN present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

'

Application Notes

Kit Components

• - Pleiotrophin (PTN Microplate (Item A)**: 96 well PCR plate coated with anti-Human Pleiotrophin (PTN

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human Pleiotrophin (PTN

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for Pleiotrophin (PTN (Item F): 2 vials of a 4x concentrated solution of anti-Human Pleiotrophin (PTN affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Canine VEGF-A IQELISA™ Kit

RayBio® Canine Immunoquantitative (PCR-Based) VEGF-A ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Canine VEGF-A IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Canine VEGF-A in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Canine VEGF-A coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and VEGF-A present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - VEGF-A Microplate (Item A)**: 96 well PCR plate coated with anti-Canine VEGF-A

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Canine VEGF-A

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for VEGF-A (Item F): 2 vials of a 4x concentrated solution of anti-Canine VEGF-A affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• Real-time PCR instrument, Bio-Rad recommended

• Precision pipettes to deliver 2 µl to 1 ml volumes

• Adjustable 1-25 ml pipettes for reagent preparation

• 100 ml and 1 liter graduated cylinders

• Absorbent paper

• Distilled or deionized water

• Log-log graph paper or computer and software for data analysis

• Tubes to prepare standard or sample dilutions

• Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human Activin R2A IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) Activin R2A ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Custom Human Activin R2A IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human Activin R2A in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human Activin R2A coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and Activin R2A present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - Activin R2A Microplate (Item A)**: 96 well PCR plate coated with anti-Human Activin R2A
• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution
• - Standards (Item C): 2 vials of recombinant Human Activin R2A
• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer
• - Detection Affinity Reagent for Activin R2A (Item F): 2 vials of a 4x concentrated solution of anti-Human Activin R2A affinity reagent
• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock
• - Primer Solution (Item I): 1.5 ml vial
• - PCR Master Mix (Item J): 1.4 ml vial
• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer
• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended
• - Precision pipettes to deliver 2 µl to 1 ml volumes
• - Adjustable 1-25 ml pipettes for reagent preparation
• - 100 ml and 1 liter graduated cylinders
• - Absorbent paper
• - Distilled or deionized water
• - Log-log graph paper or computer and software for data analysis
• - Tubes to prepare standard or sample dilutions
• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed
• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature
• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature
• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour
• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well
• 6. Run real-time PCR

Human C-type Natriuretic Peptide/CNP IQELISA™ Kit'

RayBio® Human Immunoquantitative (PCR-Based) C-type Natriuretic Peptide (CNP) ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Custom Human C-type Natriuretic Peptide (CNP IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human C-type Natriuretic Peptide (CNP in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human C-type Natriuretic Peptide (CNP coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and C-type Natriuretic Peptide (CNP present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - C-type Natriuretic Peptide (CNP Microplate (Item A)**: 96 well PCR plate coated with anti-Human C-type Natriuretic Peptide (CNP

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human C-type Natriuretic Peptide (CNP

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for C-type Natriuretic Peptide (CNP (Item F): 2 vials of a 4x concentrated solution of anti-Human C-type Natriuretic Peptide (CNP affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-26 IQELISA™ Kit'

RayBio® Human Immunoquantitative (PCR-Based) IL-26 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-26 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-26 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-26 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-26 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - IL-26 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-26

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-26

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-26 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-26 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 50 µl of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-8/CXCL8 IQELISA™ Kit

RayBio^® Human Immunoquantitative (PCR-Based) IL-8/CXCL8 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-8 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-8 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-8 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-8 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curves

Typical Data

Spiking & Recovery Results

Serum spike tests show recovery is 119% with a range of 117% to 128%

Application Notes

Kit Components

• - IL-8 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-8

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-8

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-8 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-8 affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human sTNFRII IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) TNF RII ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human TNF RII IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human TNF RII in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human TNF RII coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and TNF RII present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - TNF RII Microplate (Item A)**: 96 well PCR plate coated with anti-Human TNF RII

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human TNF RII

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for TNF RII (Item F): 2 vials of a 4x concentrated solution of anti-Human TNF RII affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).
 

Other Materials Required

• Real-time PCR instrument, Bio-Rad recommended

• Precision pipettes to deliver 2 µl to 1 ml volumes

• Adjustable 1-25 ml pipettes for reagent preparation

• 100 ml and 1 liter graduated cylinders

• Absorbent paper

• Distilled or deionized water

• Log-log graph paper or computer and software for data analysis

• Tubes to prepare standard or sample dilutions

• Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human VEGF IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) VEGF ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Qunatitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human VEGF IQELISA™ kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of human VEGF in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for human VEGF coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and VEGF present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. Ct values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower Ct values indicate a higher concentration of antigen.

Standard Curves

Typical Data

Spiking & Recovery Results

Serum spike tests show recovery is 93.3% with a range of 89.7 to 98.0%

Application Notes

Kit Components

• - VEGF Microplate (Item A)**: 96 well PCR plate coated with anti-human VEGF

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant human VEGF

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for VEGF (Item F): 2 vials of a 4x concentrated solution of anti-human VEGF affinity reagent

• - Primer Solution (Item G): 1.7 ml vial

• - PCR Master Mix (Item H): 1.2 ml vial

• - PCR Preparation buffer (Item I): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item J): 10 ml vial of 10x concentrated buffer

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 15µL Primer solution to each well

• 5. Add 10µL of PCR Master Mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Mouse IL-17A IQELISA™ Kit

RayBio® Mouse Immunoquantitative (PCR-Based) IL-17A ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Mouse IL-17A IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Mouse IL-17A in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Mouse IL-17A coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-17A present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - IL-17A Microplate (Item A)**: 96 well PCR plate coated with anti-Mouse IL-17A

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Mouse IL-17A

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-17A (Item F): 2 vials of a 4x concentrated solution of anti-Mouse IL-17A affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Rat VEGF-A IQELISA™ Kit

RayBio® Rat Immunoquantitative (PCR-Based) VEGF-A ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Rat VEGF-A IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Rat VEGF-A in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Rat VEGF-A coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and VEGF-A present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - VEGF-A Microplate (Item A)**: 96 well PCR plate coated with anti-Rat VEGF-A

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Rat VEGF-A

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for VEGF-A (Item F): 2 vials of a 4x concentrated solution of anti-Rat VEGF-A affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human CHI3L1 IQELISA™ Kit

RayBio® Human Immunoquantitative (PCR-Based) CHI3L1 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human CHI3L1 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human CHI3L1 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human CHI3L1 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and CHI3L1 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve

Typical Data

Application Notes

Kit Components

• - CHI3L1 Microplate (Item A)**: 96 well PCR plate coated with anti-Human CHI3L1

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human CHI3L1

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for CHI3L1 (Item F): 2 vials of a 4x concentrated solution of anti-Human CHI3L1 affinity reagent

• - IQELISA„™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25 µl standard or sample to each well. Incubate for 1.5 - 2.5 hours at room temperature

• 3. Add 25 µl detection antibody to each well. Incubate 1 hour at room temperature

• 4. Add 25 µl of IQELISA„™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 10 µl Primer solution and 10 µl of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2° to 8°C from the date of shipment. Standard (recombinant protein) should be stored at -20°C or -80°C (recommended at -80°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2° to 8°C. Note: the kit can be used within one year if the whole kit is stored at -20°C. Avoid repeated freeze-thaw cycles.

Human IL-2 IQELISA™ Kit

RayBio^® Human Immunoquantitative (PCR-Based) IL-2 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Human IL-2 IQELISA™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IL-2 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IL-2 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IL-2 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Standard Curves

Typical Data

Spiking & Recovery Results

Serum spike tests show recovery is 103% with a range of 109% to 97%

Application Notes

Kit Components

• - IL-2 Microplate (Item A)**: 96 well PCR plate coated with anti-Human IL-2

• - Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution

• - Standards (Item C): 2 vials of recombinant Human IL-2

• - Assay Diluent B (Item E): 15 ml of 5x concentrated buffer

• - Detection Affinity Reagent for IL-2 (Item F): 2 vials of a 4x concentrated solution of anti-Human IL-2 affinity reagent

• - IQELISA™ Detection Reagent (Item G): 1 ml of a 10x concentrated stock

• - Primer Solution (Item I): 1.5 ml vial

• - PCR Master Mix (Item J): 1.4 ml vial

• - PCR Preparation buffer (Item K): 1 ml vial of 10x concentrated buffer

• - Final Wash Buffer (Item L): 10 ml vial of 10x concentrated buffer

**The PCR plate used is a 0.2 ml, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600). Please ensure compatibility with your PCR machine prior to purchase. For additional information contact technical support (cs@medikonia.com).

Other Materials Required

• - Real-time PCR instrument, Bio-Rad recommended

• - Precision pipettes to deliver 2 µl to 1 ml volumes

• - Adjustable 1-25 ml pipettes for reagent preparation

• - 100 ml and 1 liter graduated cylinders

• - Absorbent paper

• - Distilled or deionized water

• - Log-log graph paper or computer and software for data analysis

• - Tubes to prepare standard or sample dilutions

• - Heating block or water bath capable of 80^°C

Protocol Outline

• 1. Prepare all reagents, samples and standards as instructed

• 2. Add 25µl standard or sample to each well. Incubate for 2.5 hours at room temperature or overnight at 4^°C

• 3. Add 25 µl detection affinity reagent to each well. Incubate 1 hour at room temperature

• 4. Add 25µL of IQELISA™ Detection Reagent to each well. Incubate 1 hour

• 5. Add 15µL Primer solution 10µL of PCR master mix to each well

• 6. Run real-time PCR

Storage/Stability

May be stored for up to 6 months at 2^° to 8^°C from the date of shipment. Standard (recombinant protein) should be stored at -20^°C or -80^°C (recommended at -80^°C) after reconstitution. Opened PCR plate or reagents may be stored for up to 1 month at 2^° to 8^°C. Note: the kit can be used within one year if the whole kit is stored at -20^°C. Avoid repeated freeze-thaw cycles.

Virus HIV-P24 IQELISA™ Kit

RayBio® Virus Immunoquantitative (PCR-Based) HIV-P24 ELISA Kit for cell culture supernatants, plasma, and serum samples.

Specifications

Introduction

The RayBio^® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA„™) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio^® Virus HIV-P24 IQELISA„™ Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Virus HIV-P24 in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Virus HIV-P24 coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and HIV-P24 present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. C_t values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower C_t values indicate a higher concentration of antigen.

Comparison

Standard Curve