Ab166708

Through Western blot analysis, Id2 expression levels were examined after Id2 expression plasmid transfection or knockdown in the tested cells. The detailed procedures were performed as previously described (7 (link)). Primary antibodies specifically against Id2 (ab166708; 1:1,000) were purchased from Abcam. For gel loading, the internal control comprised an anti-glyceraldehyde-3-phosphate dehydrogenase (anti-GAPDH, GT239; 1:10,000) antibody (GeneTex, Inc. Irvine, CA, USA). The membranes were first incubated with the primary antibodies. Subsequently, Tris-buffered saline and Tween 20 solution (Abcam) was applied to wash the membranes three times, after which they were incubated with horseradish peroxidase–conjugated secondary antibodies (Santa Cruz Biotechnology, Inc., Dallas, TX, USA). Signals were detected using enhanced chemiluminescence (ECL, GE Healthcare, Piscataway, NJ, USA).

Cycloheximide (CHX) and MG132 were purchased from MCE (HY-12320; MedChemExpress, China) and Selleck (S2619, China), respectively. The antibodies used in this study were as follows: anti-β-actin (M1210-2; HUABio, Hangzhou, China), anti-SMURF2 (12024S; CST, Danvers, Massachusetts, USA; bs-4056R; Bioss, Beijing, China), anti-ID2 (ab166708; Abcam, China; 3431S; CST; MA5-32891; Invitrogen, USA), anti-E2A (ab228699; Abcam), anti-Flag (14793S; CST), anti-GFP (OSE00003G; Invitrogen), anti-HA (3724T; CST), anti-Rabit IgG (3900S; CST), anti-p21 (10355-1-ap; Proteintech, Wuhan, Hubei, China; ab107099; Abcam; 2947T; CST), anti-Ubiquitin (3936T; CST).