Human IGFBP-3 HEK293 Overexpression Lysate: Product Information
This Human IGFBP-3 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of IGFBP-3 protein (Cat: 10430-H07H) from the overexpression lysate was verified.
A DNA sequence encoding the mature form of human IGFBP3 (NP_000589.2) (Gly28-Lys291) was fused with a polyhistidine tag at the N-terminus.
The secreted recombinant human IGFBP3 comprises 280 amino acids with a predicted molecular mass of 31 kDa. As a result of glycosylation, the apparent molecular mass of rhIGFBP3 is approximately 40-45 kDa in SDS-PAGE under reducing conditions.
Human IGFBP-3 HEK293 Overexpression Lysate: Usage Guide
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.
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 IGFBP-3 HEK293 Overexpression Lysate: Synonyms
Human BP-53 Overexpression Lysate; Human IBP3 Overexpression Lysate; Human IGFBP3 Overexpression Lysate
IGFBP-3 Background Information
The Insulin-like Growth Factor (IGF) signaling system plays a central role in cellular growth, differentiation, and proliferation. IGFBP3 is the most abundant IGF binding protein in human serum and is a growth inhibitory, apoptosis-inducing molecule, capable of acting via IGF-dependent and IGF-independent mechanisms. It appears to function both by cell cycle blockade and the induction of apoptosis. IGFBP3 can be transported to the nucleus by an importin beta mediated mechanism, where it has been shown to interact with the retinoid X receptor alpha and possibly other nuclear elements. IGFBP3 antiproliferative signaling appears to require an active transforming growth factor-beta (TGF-beta) signaling pathway, and IGFBP3 stimulates phosphorylation of the TGF-beta signaling intermediates Smad2 and Smad3. IGFBP3 has IGF-independent roles in inhibiting cell proliferation in cancer cell lines. Nuclear transcription factor, retinoid X receptor (RXR)-alpha, and IGFBP3 functionally interact to reduce prostate tumor growth and prostate-specific antigen in vivo. Several clinical studies have proposed that individuals with IGFBP3 levels in the upper range of normal may have a decreased risk for certain common cancers. This includes evidence of a protective effect against breast cancer, prostate cancer, colorectal cancer, and lung cancer. Moreover, IGFBP3 inhibits insulin-stimulated glucose uptake into adipocytes independent of IGF.
insulin-like growth factor binding protein 3
Baxter RC. (2001) Signalling pathways involved in antiproliferative effects of IGFBP-3: a review. Mol Pathol. 54(3): 145-8.
Butt AJ, et al. (2001) IGFBP-3 and apoptosis--a license to kill? Apoptosis. 6(3): 199-205.
Ali O, et al. (2003) Epidemiology and biology of insulin-like growth factor binding protein-3 (IGFBP-3) as an anti-cancer molecule. Horm Metab Res. 35(11-12): 726-33.
Yamada PM, et al. (2009) Perspectives in mammalian IGFBP-3 biology: local vs. systemic action. Am J Physiol Cell Physiol. 296(5): C954-76.
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