On Western blots against yeast protein, these antibodies often recognize a single cross-reacting band of about 50 or 125kD, respectively, although the level of the 50 kD polypeptide can be reduced by using cultures in the early phase of exponential division. The antibody cleanup protocol is now a seperate document Protocol Cells are grown in 5 mls of YPAD to an O.D. 600 of 0.75 to 1. The suspension is incubated at 37 o C without shaking and checked periodically.
Excess solution is aspirated.
The solution is allowed to sit for 5 minutes before aspiration. The slide is set on wet paper towels in a sealed chamber and incubated at 4 o C overnight. Excess solution is aspirated. The slide is set on wet paper towels in a sealed chamber and incubated at room temperature for 2 hours. Excess solution is aspirated.