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.

Catalog #: PEL-FYN-Y530-T-1 -- 1 Plate Kit

Catalog #: PEL-FYN-Y530-T-2 -- 2 Plate Kit

Catalog #: PEL-FYN-Y530-T-5 -- 5 Plate Kit

Please email us at cs@medikonia.com for any enquiry. To place an order, please include the catalog number(s) of the product(s) in the email.

Manual

Safety Data Sheet