Anti cyclind1 ab16663

The cells were lysed in RIPA buffer for 20 minutes at 0 °C. SDS-PAGE was used to isolate the proteins and transfer them to a PVDF membrane (BioRad). Primary antibodies (1:2001000) were used to incubate the membranes, followed by HRP-conjugated secondary antibodies. ECL plus reagents were used to detect protein bands (Bio-Rad). Primary antibodies used were anti-RSPO2 (17781-1-AP; Proteintech), anti-β-cantenin (8480S; Cell Signaling Technology), anti-phosphorylated β-catenin (Ser33/37/Thr41) (#9561, Cell Signaling Technology), anti-CyclinD1 (ab16663; Abcam) and anti-GAPDH (2188; Cell Signaling Technology).

Streptozocin (S0130) and D‐glucose (G8270) were obtained from Sigma‐Aldrich Chemical Company (Sigma). BBR (BWC9020‐2016) was procured from BeNa Biotechnology. Membrane and cytosolic protein extraction kits (P0033), trypan blue (ST798), EdU (5‐ethynyl‐2′‐deoxyuridine) assays (C0071S) and cell cycle detection kits (BB‐4104) were acquired from Beyotime Biotechnology. The 2‐NBDG (2‐deoxy‐2‐ [(7‐nitro‐2,1,3‐benzoxadiazol‐4‐yl) amino]‐D‐glucose) assay (N13195) was obtained from Thermo Fisher Scientific. Anti‐PI3K‐p85 (#4292), anti‐Akt (4691S), anti‐p‐Akt (#4060), anti‐AS160 (#2670), anti‐p‐AS160 (#8619) and anti‐GAPDH (#5174) antibodies were obtained from Cell Signaling Biotechnology. Rabbit anti‐GLUT1 (ab115730) and anti‐Cyclin D1 (ab16663) antibodies were purchased from Abcam Biotechnology (Abcam).