Anti lamin a c 4c11

A total of 1 × 10⁶ cells were lysed in 1× Cell Lysis Buffer (Cell Signaling Technology) to obtain whole-cell lysates. Equalized whole-cell lysates were used for immunoblot analysis as previously reported (60 (link)). Antibodies were used for immunoblot analysis as follows: anti–p-mTOR (Ser²⁴⁴⁸), anti-mTOR, anti-Foxo1, anti–p-Foxo1 (Ser²⁵⁶), anti–4E-BP1, anti–p-4E-BP1 (Thr^37/46), anti–p-p70S6K (Thr⁴²¹/Ser⁴²⁴), anti-p70S6K, anti–p-Stat3 (Tyr⁷⁰⁵), anti-Stat3, anti–p-Smad2 (Ser^465/467), anti-Smad2, anti–p-Stat5 (Tyr⁶⁹⁴), anti-Stat5, anti-Lamin A/C (4C11), and anti–PP2A-A were obtained from Cell Signaling Technology; anti–glyceraldehyde phosphate dehydrogenase and anti-GSTM1 were from ProteinTech; anti–β-actin was from Santa Cruz Biotechnology. The nucleus and cytoplasm proteins were extracted according to the manufacturer’s instructions by NE-PER Nuclear and Cytoplasmic Extraction Reagents (Thermo Fisher Scientific).
For immunoprecipitation, 5 × 10⁶ T cells were harvested and then lysed in phosphatase extraction buffer [20 mM imidazole-HCl, 2 mM EDTA, and 2 mM EGTA (pH 7.0)]. Foxo1 was immunoprecipitated using anti-Foxo1 antibody (Cell Signaling Technology) and protein A agarose. Beads were washed four times in 1× tris-buffered saline, and the proteins were determined by immunoblot analysis.

Cells were lysed with NETN lysis buffer (100 mM NaCl, 0.5 mM EDTA, 50 mM Tris–HCl, pH 8.0 and 0.5% NP-40) with protease inhibitor cocktail (Roche cOmplete, EDTA-free). The protein samples were separated by SDS-PAGE gel, and then transferred to nitrocellulose membrane. The following antibodies were used; anti-ATRX antibody (H-300 Santa Cruz Biotechnology, A301-045A Bethyl Laboratories), DAXX (M-112 Santa Cruz Biotechnology), Anti-Lamin A/C (4C11 Cell Signaling), and Actin (A1978 Sigma-Aldrich).