Western blot analysis of extracts from concentrated culture medium of ScaBER, U-118 MG, and SK-OV-3 cell lines using IGFBP7 Antibody (upper) and LAMC1 Antibody #92921 (lower).
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
IGFBP7 Antibody recognizes endogenous levels of total IGFBP7 protein.Species Reactivity:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human IGFBP7 protein. Antibodies are purified by protein A and peptide affinity chromatography.
IGFBP7 (also known as Mac25, TAF, or IGFBP-rP1) belongs to the IGFBP superfamily, which plays an integral role in regulating insulin-like growth factor (IGF) actions in a wide variety of cell types. There are six known high-affinity IGF binding family members (IGFBP1-6), and ten low-affinity IGF binding members. These family members are structurally related, but encoded by distinct genes (1,2). IGFBP7 is a low-affinity IGF binding protein (1). The protein functions through its binding to secreted growth factors including IGF1, insulin, and activin (3,4). IGFBP7 levels have been related to cancer development and tissue injury. Loss of expression of IGFBP7 has been associated with poor survival in multiple cancer types (5,6) and with tumor chemotherapy resistance (7,8). IGFBP7 also has been identified as a cell cycle arrest biomarker for human acute kidney injury (AKI) and serves as a prognostic indicator for early stage AKI development (9-11).
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