Benefits of Secondary Antibodies for Co-Immunoprecipitation Western Blot

This article deals with the best secondary antibodies to use when proteins that are relevant to physiological functioning are bound to the target protein by co-immunoprecipitation (co-IP). These proteins can be identified and isolated, and interactions between the proteins can be revealed and studied, in the absence of any interference from the light and heavy chains of the primary antibody after the steps of co-IP.

Common issues related to secondary antibodies for western blotting, after the co-IP step, are the signals generated due to the denatured primary antibody heavy (~50 kDa) or light chains (~25 kDa). This can result in a masking effect, which makes it difficult to detect your protein of interest if it migrates to around either the 50 kDa or 25 kDa markers.

In order to resolve this, secondary antibodies have been developed by Abcam which are optimized for co-IP, they only recognize primary antibodies in their native state. This eradicates the detection of denatured primary heavy and/or light chains during western blotting.

In addition, the secondary antibodies that have been optimized are conjugated to horseradish peroxidase (HRP), alkaline phosphatase (AP), or to biotin, making them very well-suited for western blotting.

The size of the protein of interest is an important factor to consider when it comes to selecting the correct secondary antibody that has been optimized for co-IP.

Secondary Antibodies Optimized for Co-IP for Full-Length Intact Proteins of any Size

Product name Conjugate Product code
Anti-goat IgG VeriBlot for IP secondary antibody HRP ab157532
Anti-mouse IgG VeriBlot for IP secondary antibody Biotin ab131367
Anti-mouse IgG VeriBlot for IP secondary antibody HRP ab131368
IgG VeriBlot for IP secondary antibody - bovine, goat, human, mouse, rat, rabbit, sheep AP ab131365
IgG VeriBlot for IP secondary antibody - bovine, goat, human, mouse, rat, rabbit, sheep HRP ab131366

 

​Light Chain-Specific Secondary Antibodies for Proteins of Interest, Greater than 30 kDa

Product name Conjugate Product code
Rat anti-mouse kappa light chain AP ab99616
Rat anti-mouse lambda light chain AP ab99624
Rat anti-mouse kappa light chain AP ab99631
Mouse anti-rabbit IgG light chain AP ab99696
Mouse anti-rat kappa light chain AP ab99691
Mouse anti-human lambda light chain HRP ab99811
Rat anti-mouse kappa light chain HRP ab99617
Rat anti-mouse lambda light chain HRP ab99632
Mouse anti-rabbit IgG light chain HRP ab99697
Mouse anti-rat kappa light chain HRP ab99692

 

About Abcam

Abcam is a global life sciences company providing highly validated antibodies and other binders and assays to the research and clinical communities to help advance the understanding of biology and causes of disease.

Abcam’s mission is to serve life scientists to help them achieve their mission faster by listening to their needs, continuously innovating and improving and by giving them the tools, data and experience they want. Abcam’s ambition is to become the most influential life science company for researchers worldwide.


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Last updated: Jun 13, 2019 at 7:56 AM

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