Anti lamin b1 d4q4z

Human wild type (Wt), N-terminal myristoylated (myr)-tagged, and C-terminal NLS-tagged PTEN complementary DNA (cDNA) sequences were subcloned from pCDNA3-PTEN into pTRIPZ vector (Addgene, Watertown, MA, USA) with AgeI and MluI to generate PTEN expression plasmid. All plasmid constructs were confirmed by sequencing. Ampicillin, puromycin, and doxycycline were purchased from Sigma-Aldrich (St. Louis, MO, USA). Polyethylenimine (PEI) was purchased from Polysciences (Warrington, PA, USA), while RPMI, DMEM, Opti-MEM reduced serum media, and fetal bovine serum (FBS) were purchased from Thermo Fisher Scientific (Waltham, MA, USA), and Protein G Sepharose 4 Fast Flow beads were purchased from Cytiva (Malborough, MA, USA). The antibodies for Western blotting were as follows: anti-PTEN (138G6, 1:1000), anti-p-AKT (9271, 1:1000), anti-AKT (9272, 1:1000), anti-GAPDH (D16H11, 1:4000), anti-Lamin B1 (D4Q4Z, 1:1000), anti-β-Tubulin (9F3, 1:1000), and anti-EGFR (D3871, 1:1000) antibodies, all purchased from Cell Signaling Technology (Danvers, MA, USA). Anti-HSP90 (610418, 1:4000) was purchased from BD Bioscience (Franklin Lakes, NJ, USA).

Cytoplasmic and nuclear proteins were separated using NE‐PER Nuclear and Cytoplasmic Extraction Reagents kits (#78833, Thermo). Cell lysates were examined by western blot analysis using the Pierce BCA Protein Assay Kit (#23225, Thermo Scientific). The following primary antibodies were purchased from Cell Signaling Technology: anti‐NFAT1 (D43B1), anti‐Lamin B1 (D4Q4Z), and anti‐β‐Actin (D6A8) antibodies.