Antibody-based Protein Biochemical Analysis

For immunolabeling and protein biochemical methods, the following antibodies have been used: anti-TRIM32-3150 (Gramsch Laboratories), anti-TRIM32-3149 (Gramsch Laboratories), anti-TRIM32-M09 (Abnova), anti-TRIM32-GS (Genescript), anti-TRIM32-1137 (Gramsch Laboratories) [13 (link), 18 (link)–20 (link)], anti-USP7 (Novus Biologicals), anti-c-Myc N-262 (Santa Cruz), anti-mono-and-polyubiquitinylated conjugates (FK2) (Enzo Life Sciences), anti-FLAG M2 (Sigma), anti-enhanced green fluorescent protein (anti-EGFP) (Abcam), anti-α-Tubulin (Sigma-Aldrich), anti-Ki67 (BD Biosciences), anti-Mash1 (BD Biosciences), anti-Nestin (BD Biosciences), anti-Tuj1 (BioLegend), anti-Doublecortin X (Millipore), anti-Neuronal Nuclei (anti-NeuN) (Millipore) and anti-GFAP (Millipore). As secondary antibodies Alexa-fluorophore-conjugated antibodies (Invitrogen) were used for immunofluorescence stainings and horseradish peroxidase-coupled antibodies (GE Healthcare) for immunoblotting. DNA was counterstained using Hoechst 33258 (Invitrogen).
The following plasmids were used: pEGFP-N1 (Clontech Laboratories), pcDNA3-GFP-TRIM32 (kindly provided by Dr. Germana Meroni), pEGFP-TRIM32-ΔRING [14 (link)], pcDNA3-c-Myc (kindly provided by Dr. Martin Eilers), pBSK-HA-Ubiquitin (kindly provided by Dirk Bohrmann), pQFlag-USP7-WT-puroR and pQFlag-USP7-CS-puroR (kindly provided by Dr. Goedele Maertens).