Mouse anti lamin a c

The following antibodies were used for western blotting: rabbit anti-SIRT2 (1:1000; Sigma #S8447 or Cell Signaling #12650), rabbit anti-ANKLE2 (1:500; Atlas Antibodies #HPA003602), goat anti-RTN4 (1:1000; Nogo N18, Santa Cruz Biotechnology #sc-11027), anti-FLAG M2-peroxidase clone M2 (1:10,000; Sigma #A8592), streptavidin–HRP (1:1000; GE Healthcare #RPN1231), mouse anti-HA.11 clone 16B12 (1:1000; Covance #MMS-101R), mouse anti-Myc clone 4A6 (1:1000; Merck Millipore #05-724), rabbit anti-acetylated-lysine (1:1000; Cell Signaling #9441), mouse anti-α-tubulin clone B512 (1:5000; Sigma #T5168), mouse anti-His (1:5000; GE Healthcare #27471001), mouse anti-GFP (1:1000; Roche #11814460001). The following antibodies were used for immunofluorescence: rabbit anti-SIRT2 (1:200; Sigma #S8447), mouse anti-Myc clone 4A6 (1:500; Merck Millipore #05-724), streptavidin–Alexa-Fluor-568 (1:500; Life Technologies #S11226), rabbit anti-calnexin (1:200; a gift from Erwin Ivessa; Medical University of Vienna, Austria), mouse anti-lamin A/C (1:100; Millipore #MAB3211), mouse anti-acetylated tubulin (1:1000; Sigma #T7451). Secondary HRP-conjugated antibodies for western blotting (Jackson ImmunoResearch) were used at 1:10,000 dilution. Secondary Alexa-Fluor^®-conjugated antibodies for immunofluorescence (Life Technologies) were used at 1:500 dilution.

Whole-cell lysates of MSC were collected, separated by SDS polyacrylamide gel electrophoresis, and transferred onto polyvinylidene fluoride membrane using the liquid transfer method. The blots were blocked in 10% skim milk (Bio-Rad, Hercules, CA, USA) in Tween 0.1% tris-buffered saline 1× 1 h at room temperature. The primary antibodies were a mouse anti-lamin A/C 1:200 (Millipore, Billerica, MA, USA; JOL2, MAB3211), a mouse anti-SFRS1/SF2 1:500 (LSBio, LS-B2340, Seattle, WA, USA) and a β-actin 1/200,000 (Sigma). The membranes were incubated during the night at 4 °C. Antigen–antibody binding was detected using horseradish peroxidase-conjugated species-specific secondary antibodies (GE-Healthcare, Little Chalfont, UK), followed by enhanced chemiluminescence western blotting detection reagents (Perkin-Elmer, Waltham, MA, USA). The western blot results were quantified using ImageJ software.

The following reagents were purchased from the indicated providers: dimethyl sulfoxide (DMSO; Sigma, D8418) and mifepristone (RU-486; Sigma, M8046). The following antibodies were used for immunoblotting: mouse anti-TurboGFP (Origene, TA150041), rabbit anti-TDP-43 (Proteintech, 10782-2-AP), rabbit anti-LC3 (MBL, PM036), mouse anti-Polyubiquitin (Enzo Life Science, BML-PW8805), mouse anti-Flag (Cell Signaling Technology, 2044), rabbit anti-HDAC6 (Santa Cruz Biotechnology, sc-11420), mouse anti-Lamin A/C (EMD Millipore, 05-714), HRP-conjugated anti-alpha-tubulin (Cell Signaling Technology, 9099), HRP-conjugated anti-rabbit IgG (Santa Cruz Biotechnology, sc-2004), HRP-conjugated mouse IgM (Abcam, ab97230), and HRP-conjugated mouse IgG (Santa Cruz Biotechnology, sc-2005). The following antibodies were used for immunocytochemistry (ICC): rabbit anti-cleaved caspase-3 (CC3) antibody (Cell Signaling Technology, 9664) and Alexa 594-conjugated anti-rabbit IgG (Jackson ImmunoResearch, 111-585-144). The following antibodies were used for immunohistochemistry: rat anti-ELAV (DSHB, RAT-ELAV-7), mouse anti-Polyubiquitin (Enzo Life Science, BML-PW8805), Alexa-488 conjugated rat IgG (Jackson ImmunoResearch, 112-545-167), and Alexa-594 conjugated mouse IgM (Jackson ImmunoResearch, 115-587-020).