A simplified 'coupled' probing approach for the detection of proteins on dot and western blots.
Western blotting is one of the most commonly and widely used methods for identifying the presence of an antigen in biochemical studies. The present study describes a novel 'coupled blotting' approach that allows antigen on dot and Western blots to be simultaneously probed with primary and secondary probing reagents. The study utilized the highly sensitive enhanced chemiluminescence (ECL) detection system, purified E7 protein of cottontail rabbit papillomavirus (CRPV) and E7-specific antibodies. Comparison of the abilities of sequential primary antibody followed by secondary reagent and coupled treatments to detect E7 protein on blots showed no reduction in signal strength after coupled probing. The new 'coupled' probing has the advantage of saving hands-on time and buffer solutions over the standard procedure. Overall, the results suggest that the 'coupled blotting' is useful for the rapid detection of proteins without compromising quality.
Sundaram
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Section of Comparative Medicine, Yale University School of Medicine, New Haven, CT 06520-8016, USA.
D000818:Animals; D015153:Blotting, Western; D012777:Cottontail rabbit papillomavirus; D015151:Immunoblotting / Q000379:methods*; D009856:Oncogene Proteins, Viral / Q000032:analysis*; D011817:Rabbits
C079183:E7 protein, cottontail rabbit papillomavirus; D009856:Oncogene Proteins, Viral