Improving the quality of immunoblots by chromatography of polyclonal antisera on keratin affinity columns - Girault, J.-A.; Gorelick, F.S.; Greengard, P. (Elsevier)
Unwanted reactivity of polyclonal antisera against keratins (''fingerprint proteins'') is a problem commonly encountered when proteins transferred to nitrocellulose are studied by immunoblotting. Immunoreactivity against keratins is generally accompanied by a spotted background. This antikeratin immunoreactivity could be removed by adsorption of the antisera to human keratin bound to nitrocellulose. Larger amounts of antisera were purified from contaminant antikeratin antibodies by a single passage over a column of human keratin coupled to activated CH-Sepharose 4B. In contrast to nonpurified antisera and their IgG fractions, the column effluent no longer recognized the M"r 55,000-70,000 keratin proteins and exhibited a marked decrease in background labeling. We propose this simple method as a valuable alternative when affinity purification of polyclonal antisera on antigen columns is not practical.
http://dx.doi.org/10.1016/0003-2697(89)90741-0
Keywords: W; V