The proliferation of HUVEC treated with increasing concentrations of hVEGF121 was assessed. After 72-hour treatment with hVEGF121 cells were incubated with a tetrazolium salt and the OD450 - OD650 was determined.
The purity of recombinant hVEGF121 was determined by SDS-PAGE of 6 µg reduced (+) and non-reduced (-) recombinant hVEGF121 and staining overnight with Coomassie Blue.
Western blot analysis of extracts from HUVEC untreated or treated with hVEGF121 for 15 minutes, using Phospho-p38 MAPK (Thr180/Tyr182) (3D7) Rabbit mAb #9215 (upper) and p38 MAPK Antibody #9212 (lower).
With carrier: Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.2 containing 20 μg BSA per 1 μg hVEGF121. Carrier free: Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.2.
Stable in lyophilized state at 4°C for 1 year after receipt. Sterile stock solutions reconstituted with carrier protein are stable at 4°C for 2 months and at -20°C for 6 months. Avoid repeated freeze-thaw cycles.Maintain sterility. Storage at -20°C should be in a manual defrost freezer.
>98% as determined by SDS-PAGE of 6 μg reduced (+) and non-reduced (-) recombinant hVEGF121. All lots are greater than 98% pure.
Recombinant hVEGF121 contains no "tags" and the nonglycosylated protein has a calculated MW of 14,057. DTT-reduced protein migrates as a 14-22 kDa polypeptide. Heterogeneity in SDS-PAGE is due to glycosylation. The non-reduced cystine-linked homodimer migrates as a 30-36 kDa protein. The expected amino-terminal APMAE of recombinant hVEGF121 was verified by amino acid sequencing.
The bioactivity of recombinant hVEGF121 was determined in a cell proliferation assay using HUVEC. The ED50 of each lot is between 0.5-2 ng/ml.
Less than 0.01 ng endotoxin/1μg hVEGF121.
Recombinant human VEGF121 (hVEGF121) Ala207-Arg327 (Accession #NP_001020541.2) was expressed in human 293 cells at Cell Signaling Technology.
VEGF121 is the second most abundant splice variant of VEGF-A (1,2). VEGF121 is produced by endothelial cells, macrophages, T-cells and other cell types. VEGF121 is involved in angiogenesis, vascular endothelial cell survival, growth, migration and vascular permeability (1). VEGF121 expression is induced by hypoxia, inflammatory cytokines and through oncogene products in tumors (1-3). VEGF121 binds to VEGFR1 and VEGFR2 receptor tyrosine kinases (1). Binding of VEGF121 to VEGFR1 and VEGFR2 leads to activation of pathways involving PI3K/AKT, P38 MAPK, and FAK (1). VEGF plays a key role in tumor angiogenesis in many cancers (2).
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