Immunoprecipitation of p27^Kip1 following denaturation with 8 M urea.

Prior to immunoprecipitation with the polyclonal anti-p27^Kip1 (C-19), the protein extracts were either treated with 8 M urea and subsequently diluted 1:20 as described in “Materials and methods” (lane 8M) or treated by dilution with buffer not containing urea (N), or diluted with buffer in a final concentration of 0.4 M urea (lane 0.4M). The p27^Kip1 antibody immunoprecipitates significantly more p27^Kip1 protein following denaturation of cell lysates with 8 M urea. Note that there is practically no difference in the expression of p27^Kip1 between the extracts with native conditions and the one treated with 0.4 M urea.