This Human FGFR1 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of FGFR1 protein (Cat: 10616-H03H) from the overexpression lysate was verified.
A DNA sequence encoding the human FGFR1 (NP_075594.1) extracellular domain (Met 1-Glu 285) was fused with the C-terminal polyhistidine-tagged Fc region of human IgG1 at the C-terminus.
The recombinant human FGFR1/Fc is a disulfide-linked homodimer after removal of the signal peptide. The reduced monomer consists of 512 amino acids and has a predicted molecular mass of 57.5 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rhFGFR1/Fc monomer is approximately 100-110 kDa due to glycosylation.
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.
Human bFGF-R-1 Overexpression Lysate;Human BFGFR Overexpression Lysate;Human CD331 Overexpression Lysate;Human CEK Overexpression Lysate;Human FGFBR Overexpression Lysate;Human FGFR-1 Overexpression Lysate;Human FLG Overexpression Lysate;Human FLT-2 Overexpression Lysate;Human FLT2 Overexpression Lysate;Human HBGFR Overexpression Lysate;Human HH2 Overexpression Lysate;Human HRTFDS Overexpression Lysate;Human KAL2 Overexpression Lysate;Human N-SAM Overexpression Lysate;Human OGD Overexpression Lysate
FGFR1, also known as CD331, belongs to the fibroblast growth factor receptor subfamily where amino acid sequence is highly conserved between members and throughout evolution. FGFR family members differ from one another in their ligand affinities and tissue distribution. Fibroblast growth factors (FGFs) (FGF1 - 10 and 16 - 23) are mitogenic signaling molecules that have roles in angiogenesis, wound healing, cell migration, neural outgrowth and embryonic development. FGFs bind heparan sulfate glycosaminoglycans, which facilitates dimerization (activation) of FGF receptors. FGFR1 is a full-length representative protein consists of an extracellular region, composed of three immunoglobulin-like domains, a single hydrophobic membrane-spanning segment and a cytoplasmic tyrosine kinase domain. The extracellular portion of FGFR1 interacts with fibroblast growth factors, setting in motion a cascade of downstream signals, ultimately influencing mitogenesis and differentiation. This particular family member binds both acidic and basic fibroblast growth factors and is involved in limb induction. CD331 can be detected in astrocytoma, neuroblastoma and adrenal cortex cell lines. Some isoforms are detected in foreskin fibroblast cell lines, however isoform 17, isoform 18 and isoform 19 are not detected in these cells. Defects in FGFR1 are a cause of Pfeiffer syndrome ，idiopathic hypogonadotropic hypogonadism， Kallmann syndrome type 2, osteoglophonic dysplasia and trigonocephaly non-syndromic.
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Schlessinger J, et al. (2000) Crystal structure of a ternary FGF-FGFR-heparin complex reveals a dual role for heparin in FGFR binding and dimerization. Mol Cell. 6(3):743-50. Dodé C, et al. (2007) Novel FGFR1 sequence variants in Kallmann syndrome, and genetic evidence that the FGFR1c isoform is required in olfactory bulb and palate morphogenesis. Hum Mutat. 28(1): 97-8. Kim HG, et al. (2005) Hypogonadotropic hypogonadism and cleft lip and palate caused by a balanced translocation producing haploinsufficiency for FGFR1. J Med Genet. 42(8):666-72.