Anti ints7 antibody

Co-IP assay was performed by incubating cell lysates with an anti-BAG3 antibody (Proteintech, Chicago, IL, USA) or an anti-INTS7 antibody (Proteintech), as previously described (Xue et al., 2022 (link)). Briefly, the cells were lysed on ice for 30 min using RIPA lysis buffer (Beyotime, Nantong, China) and then precleared with protein A agarose(Invitrogen). The lysates were incubated with the indicated antibodies overnight at 4 °C and then with protein A agarose at 4 °C for 3 h. The immunoprecipitants were washed with PBS, eluted with sodium dodecyl sulfate buffer, and analyzed by western blotting.

Briefly, 5 μg of anti-INTS7 antibody (Proteintech) was incubated with total BM-MSC lysates for 2 h, followed by the addition of 20 μl of washed protein A/G beads (Santa Cruz Biotechnology) and incubated for another 16 h at 4°C. Beads were then pelleted and washed three times in 20× bed volume of lysis buffer. The bound protein was eluted by heating the beads at 95°C for 5 min with 2× SDS buffer. Finally, anti-INTS7 (Proteintech), anti-ABCD3 (Abcam), or anti-HDLBP (Proteintech) antibodies were used to detect the presence of each protein in the immunoprecipitation (IP) product. Inputs represented approximately 1/10 of the extract volume used for the IP experiment.