Anti pfgfr1

A commercially available tissue microarray (TMA) (CNS2081, US Biomax) with 208 cores obtained from CNS pathologies was screened for FGFR1 and pFGFR1 expression by immunohistochemistry (IHC). The TMA included adult and pediatric HGGs, LGGs, meningiomas, and medulloblastomas alongside healthy controls. Prior to staining, the TMA slide was heated at 70°C before immersion in xylene. Rehydration in absolute ethanol (Sigma) was followed by 90% ethanol and finally 70% ethanol. Enzyme antigen retrieval was achieved using Bond Enzyme Pre-treatment Kit (Leica Biosystems). Endogenous peroxidase activity was blocked using Bloxall (Vector Laboratories). Protein blocking was then performed with 10x casein (1:10) (Vector Laboratories). Rabbit anti-FGFR1 primary antibody (Sigma) or anti pFGFR1 (Cell signaling) was added at 1:3,000 and 1:100 concentrations followed by incubation with the anti-rabbit polymer horseradish peroxidase (HRP) secondary antibody (MenaPath). The slide was developed with ImmPACT 3,3'-diaminobenzidine (DAB) HRP substrate (Vector Laboratories) and counterstained using in-house Mayer's haematoxylin, dehydrated and mounted with DPX. An Aperio scanner (Leica Microsystems, Milton Keynes, UK) was used for visualization and scoring of the stained cores.

WZ4002 (#S1173) and AZD4547 (#S2801) were purchased from Selleck Chemicals. Human bronchial epithelial cell line BEAS-2B, human lung squamous cancer cell line H520, and human NSCLC cell lines H1975 and PC9 were procured from the Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences and tested for mycoplasma contamination by DAPI staining before experiment. All the cells were maintained in RPMI-1640 medium (#C11875500BT, Gibco) with 10% FBS (#10270-106, Gibco), 100 μg/ml streptomycin, and 100 U/ml penicillin (#15140122, Gibco) and placed in a humidified cell incubator (5% CO₂, 37°C). Antibodies including anti-p-EGFR (#3777S), anti-p-FGFR1 (#2544S), anti-EGFR (#2646S), anti-FGFR (#9740S), anti-GAPDH (#5174S), and HRP-linked anti-rabbit IgG (#7074S) were purchased from Cell Signaling Technology (Danvers, MA, USA).