Secondary anti mouse igg hrp

The cell medium was centrifuged at 4 °C for 10 min at 12,000 rpm to remove cells and debris, and the supernatant was collected and stored at -80 °C for further analysis. The transfected cells were harvested and treated with RIPA lysis buffer (Beyotime, China) for 30 min. The cell suspension was centrifuged at 4 °C for 10 min at 12,000 rpm, while the lysates and supernatants were collected and stored at -80 °C. Western blotting was performed to analyze the expression of LPL protein in both the cell medium and lysate. The antibodies (all from Santa Cruz Biotechnology (Shanghai)) and antibody dilutions used in this study were as follows: primary rabbit LPL antibody (product number 73,646), 1:200; primary mouse GAPDH antibody (product number 47,724), 1:5000; secondary anti-rabbit IgG-HRP (product number 2357), 1:2000; and secondary anti-mouse IgG-HRP (product number 2004), 1:5000. Analysis of band intensity was performed by means of ImageJ software. Analysis of LPL activity in the cell medium was performed as previously described [29 (link)]. All experiments were repeated at least 3 times independently. The results of Western blotting and LPL activity analysis are shown as the mean ± standard deviation (SD) and were analyzed by the SPSS 25.0 software package (IBM Analytics, Armonk, NY). A probability (P value) of less than 0.05 was defined as being statistically significant.

CXCR4 antibody was used at a concentration of 1ug/ml (Cat#ab2074, Abcam, Cambridge, MA 02139) and secondary donkey anti-rabbit IgG HRP at 1:5000 dilution (Cat#sc-2313, Santa Cruz Biotechnology, Santa Cruz, CA). β-actin (Cat#sc-47778) antibody was used at 1:500 dilution and secondary anti-mouse IgG HRP (Cat#sc-2314) at 1:2000 dilution (Santa Cruz Biotechnology, Santa Cruz, CA). β-actin expression was used to confirm equal protein loading on the SDS-PAGE gels.