Anti c abl a5844

Proteins were prepared as described previously (Cancino et al., 2008 (link)). Tissue protein samples (30 μg) and cellular protein samples (50 μg) were resolved by SDS–PAGE. The immunoblot was carried out using anti-c-Abl (A5844, Sigma-Aldrich, USA), p-c-Abl (Tyr412) (07–788; Millipore), anti-LC3 (NB100-2220), anti-p62 (ab56416), and anti-GAPDH (0411; sc47724; Santa Cruz Biotechnology) antibodies. The secondary antibodies against rabbit or mouse IgGs conjugated with horseradish peroxidase were obtained from Upstate Biotechnology, Lake Placid, NY, United States.

Hippocampal neurons from WT and c-Abl-KO embryos were plated at a density of 10⁶ cells/cm². At different DIV, they were washed and lysed in radioimmunoprecipitation assay (RIPA) buffer (50 mM Tris, 150 mM NaCl, 1 mM EGTA, 1 mM EDTA, 0.5% deoxycholate, 1% NP-40, and 0.1% SDS) supplemented with protease inhibitors. Cell lysates were centrifuged at 14,000 rpm for 15 min at 4°C. Protein quantification was performed using the Pierce^® BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA). The fractions were subjected to SDS-PAGE and transferred onto nitrocellulose membranes (Thermo Fisher Scientific, Waltham, MA, USA). The antibodies used were: anti-βIII-tubulin (AA10 sc80016, Santa Cruz Biotechnology, Dallas, TX, USA), anti-c-Abl (A5844, Sigma-Aldrich, St. Louis, MO, USA); anti-PSD95 (75–028) and anti-SAP102 (75–058), from NeuroMAb Antibodies Inc.