The Ins and Outs of Antibody Stability

What Makes Antibodies Unstable? Antibody molecules are prone to instability during storage whether at room temperature or even at -200C. Oxidation and proteolytic degradation of proteins are significant concerns for antibody stability even at 40C and the frequency of these reactions significantly increases at higher temperatures. Another concern for antibody stability is created when antibodies […]

All Purified Antibodies are NOT Created Equal

One of the key issues with polyclonal antibody quality is the purity of the antibody preparation. Polyclonal antibodies are sold in many different preparations including “crude serum” and “purified”. Crude serum has an IgG concentration of about 10 mg/ml, but the concentration of any specific antibody in that pool of IgG is rarely more than […]

Why do so many researchers prefer polyclonal antibodies?

In an article in Nature on antibody quality (Monya Baker, Nature  527:545-551, 2015) a study was cited in which antibodies were used in over 2000 studies.  Among these polyclonal antibodies were used in 1293 studies, monoclonals were used in 755 papers.  This preference for polyclonal antibodies has been echoed in a number of other more […]

Secondary Antibody Issues are Often Overlooked in Immunochemistry

The Choice of Secondary Antibody is one of the Most Overlooked Issues in Immunochemistry. After the tissue of interest has been incubated with the primary antibody, the next critical step involves incubating the tissue in a labeled secondary antibody to detect the presence of the primary antibody. The choice of the secondary antibody has to […]

Why Don’t More Researchers Validate Antibodies with WB Before Doing IHC?

  As we have discussed previously in this blog, there is widespread concern about antibody validation. Of course some forms of validation using multiple antibodies or knockouts are not available for the majority of targets of interest to researchers.  But western blots are a excellent form of validation that can be readily performed by almost […]