Anti hmga2

Post‐treatment with different doses of metformin, total protein of MCF‐7 and BT‐474 cells were extracted by using RIPA lysis buffer. The same amount of total protein for each sample was running in SDS‐PAGE gel. The primary antibodies including anti‐HMGA2 and anti‐β‐actin were purchased from GeneTex Inc. (Irvine, USA) and Proteintech (Chicago, USA). The secondary antibodies including goat anti‐rabbit (Abcam, USA) or anti‐mouse antibody (Abcam) were incubated with western blots and the membrane was then visualized by ECL (Millipore). Image J software was used to quantify the intensity in western blotting analysis.

Anti-hLin41 (R&D), anti-Lin28B (Cell Signaling Technology), anti-Lin28A (Abcam), anti-β-actin (clone C4, Millipore), anti-α-tubulin (clone TU-02, Santa Cruz Biotech), anti-CPSF73 (Bethyl Laboratories), anti-symplekin (BD Biosciences), anti-HuR (Santa Cruz Biotech), anti-Ras (Abcam), anti-eIF4GI [6 (link)], anti-FLAG (clone L5, BioLegend), and anti-HMGA2 (GeneTex) antibodies were used through the all western blot analysis.

The single-cell suspension was obtained by mechanical disaggregation of the spheres and total proteins were extracted 48 h later, as previously described^{38 (link)}. After separation by SDS–polyacrylamide gel electrophoresis, proteins were blotted on nitrocellulose membranes (GE Healthcare Europe Gmb) and hybridized with the following antibodies: anti-HMGA1, anti-HMGA2 (Genetex) or affinity-purified anti-HMGA2, anti-α-actin (SantaCruz Biotechnologies).