DESCRIPTION

FGFR2 (Fibroblast growth factor receptor 2) is a tyrosine-protein kinase that acts as a cell-surface receptor for fibroblast growth factors and plays an essential role in the regulation of cell proliferation, differentiation, migration, and apoptosis. Ligand binding leads to the activation of several signaling pathways, such as RAS, MAPK1/ERK2, MAPK3/ERK1, and the MAP Kinase signaling pathway, as well as the AKT1 signaling pathway. Mutations in the gene of FGFR2 are associated with Crouzon syndrome, Pfeiffer syndrome, Craniosynostosis, Apert syndrome, Jackson-Weiss syndrome, Beare-Stevenson cutis gyrata syndrome, Saethre-Chotzen syndrome, and syndromic craniosynostosis.

PRINCIPLE OF THE ASSAY

An anti-tag antibody is pre-coated onto the bottom of wells. After adding antigen or samples, Tagged-Capture antibody and HRP-conjugated detection antibody, a sandwich complex is formed in the solution. TMB acts as a HRP substrate, and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns the solution yellow. The color intensity is proportional to the quantity of bound protein, which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 8 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 16 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	8	4,419.1	52.5	1.2
2	8	994.5	48.7	4.9
3	8	530.7	24.1	4.5

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	4,424.6	58.8	1.3
2	16	998.2	43.5	4.4
3	16	531.0	23.6	4.4

RECOVERY

The recovery of human FGFR2 spiked to three different levels throughout the range of the assay in various samples was evaluated.

Sample Type		Average% of Expected	Range (%)
Human plasma	1:8	99	94-104
	1:16	95	87-100
Cell lysate	1:2	91	85-101
	1:4	93	92-95

LINEARITY

To assess the linearity of the assay, cell lysate samples were spiked with high concenthumanions of human FGFR2 and diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay. Human plasma samples were diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay.

		Human plasma(Sample Diluent PT 5)	Cell lysate(Sample Diluent PT 5-ec)
1:2	Average% of Expected	100	89
	Range (%)	-	87-91
1:4	Average% of Expected	95	87
	Range (%)	91-100	85-90
1:8	Average% of Expected	91	89
	Range (%)	78-104	88-90
1:16	Average% of Expected	107	88
	Range (%)	103-111	87-89

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human FGFR2 produced at Sunlong Medical™.