Study reveals how treatment-resistant prostatecancer provides its own hormonal fuel wustl NatureComms
Two rabbits were immunized twice with the acetyl peptide, and the sera from these rabbits was affinity purified. Two antigen-affinity columns were used to purify the acetyl-specific antibodies. The first column was the non-acetyl peptide affinity column. Antibodies recognizing the non-acetylated residues of the peptide bound to the column. The flow-through fraction was collected and then applied to the second column, the acetyl peptide column.
For immunoprecipitation studies, cell lysed by sonication in RLB, the lysates were quantitated, and 0.5 to 1 mg of protein lysate was immunoprecipitated using 2 μg of anti-SREBF1, anti-pTyr, anti-FLAG, anti-HA, anti-Myc or anti-AR antibody coupled with protein A/G sepharose overnight, followed by washes with RLB and PBS buffers. The beads were boiled in a sample buffer and immunoblotting was performed as described above.
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