2.7 Co-immunoprecipitation (Co-IP) assay
2×106 293T cells were seeded in 10 cm culture dishes
and transfected 12 h later. Forty-eight hours post-transfection, the
cells were lysed with RIPA buffer and cleared by centrifugation. The
supernatants were collected and incubated with anti-FLAG
antibody-conjugated magnetic beads (Sigma) overnight at 4°C. Then, the
beads were collected with a magnetic rack, repeatedly washed, and used
for the WB assay.