Phospho-FGFR2 (Tyr653/654) & Total FGFR2 Cellular Assay Kits

Phospho- and total FGFR2 assays for the pharmacological study of FGF binding in cancer and CNS research

FGFR2 (also called CD332) is a receptor for FGF (fibroblast growth factor). Like all growth factor receptors, ligand binding induces receptor dimerization / autophosphorylation ans subsequent signaling through MAPK and Pi3K/Akt pathways. Mutations in FGFR2 have been associated with cancer (breast, melanoma) and abnormal bone development (cranosynostosis syndromes).

Our offer includes cell-based phospho FGFR2 (Tyr653/654) assay to monitor receptor activation and total FGFR2 to normalize against steady state expression.

   Check our lysis buffer compatibility      Check our species compatibility

Assay principle

HTRF® - the homogeneous cell-based sandwich immunoassay

Cisbio Bioassay’s phospho- and total FGFR2 assays are based on a TR-FRET sandwich immunoassay format comprising two specific anti-FGFR2 antibodies, one labeled with Eu3+-cryptate (donor) and the other with d2 (acceptor). The antibodies specifically bind with FGFR2, and the proximity of donor and acceptor will then lead to a fluorescent TR-FRET signal. The protocol is optimized for a 384-well plate format, but can easily be further miniaturized or upscaled. Only low sample volumes are needed. The detection reagents may be pre-mixed and added in a single dispensing step for direct detection. No washing is needed at any step.
The assays can be run with frozen cell lysates or fresh cells in culture. After cell lysis, endogenous phospho- or total-FGFR2 can be quantitatively detected using the HTRF phospho-FGFR2 (Tyr653/654) and total FGFR2 cellular assay kit reagents and most TR-FRET multimode plate readers.

A simpler, more flexible assay protocol - adapted to your applications

Two-plate assay protocol

For added flexibility, the assay can be run under a two-plate assay protocol, where cells are plated and treated in a 96-well culture plate. For detection, lysates are subsequently transferred to a 384sv assay plate where the HTRF® reagents are added. This also enables monitoring of the cells' viability and confluence in an appropriate cell culture plate.

What to expect at the bench

This video covers the principles of phospho-HTRF assays and gives guidelines for performing them, using our phospho-ERK assay as an example.

Product Performance

1. Detection phospho and total FGFR2 on Kato-III cells

Kato-III cells were plated in 96-well plate and incubated overnight. Cells were treated with increasing concentration of FIIN-1 hydrochloride compound for 6h at 37 °C - 5% CO2. After stimulation, cells were washed and lysed with lysis buffer#2 1X for 30 min at RT under gentle shaking. 16µL of lysate were transferred into a 384-well sv white microplate and 4 µL of the HTRF phospho FGFR2 or total FGFR2 detection reagents were added. The HTRF signal was recorded after a overnight incubation at room temperature.

Part#, inserts & MSDS

Ordering Info

DescriptionCat. noProduct insertMSDS
FGFR2 phospho-Y653/654 kit - 500 tests63ADK054PEG
FGFR2 phospho-Y653/654 kit - 10,000 tests63ADK054PEH
FGFR2 phospho-Y653/654 kit - 50,000 tests63ADK054PEY
FGFR2 total kit - 500 tests63ADK057PEG
FGFR2 total kit - 10,000 tests63ADK057PEH
FGFR2 total kit - 50,000 tests63ADK057PEY

Companion products

DescriptionCat. noProduct insertMSDS
FGFR2 phospho-Y653/654 kit - control lysate63ADK054TDA
FGFR2 total kit - control lysate63ADK057TDA
Phospho-total protein blocking reagent - 2 ml64KB1AAC
Phospho-total protein blocking reagent - 6 ml64KB1AAD