Anti rabbit hrp antibodies

Cell protein extracts (10 μg) mixed with the DTT-containing Laemmli loading buffer were incubated at 40 °C for 10 min, applied to 8% SDS-PAGE, and then transferred onto the nitrocellulose membrane (GE Healthcare) in 25 mM Tris, 192 mM glycine, and 20% methanol, with subsequent membrane blocking in 5% nonfat milk in PBS for 1 h at room temperature. Antibodies against total OCT-1, OCT-1A isoform, Lamin B, or beta-actin were used in the Western blot assay. Lamin B or beta-actin was used as a loading control for subsequent normalization. Membranes were probed with primary antibodies in PBS supplemented with 0.1% Tween-20 (PBS-T) overnight and then washed 3 times for 20 min with PBS-T and incubated for 1 h at RT with the anti-mouse HRP antibodies (Santa-Cruz Biotechnology, sc-2005, 1:5000) or anti-rabbit HRP antibodies (Santa-Cruz Biotechnology, sc-2005, 1:5.000). After four washing steps with PBS-T, signal detection was performed according to the standard protocol using the ECL-reagent (GE Helthcare). Western blot assay results were visualized, and the obtained signal was quantitatively assessed using the ChemiDoc MP Imaging System (Bio-Rad) with the aid of the Bio-Rad Image Lab software. In each experiment, the measurements were made in triplicates, and mean values were calculated.

Neuronal nuclei (NeuN), Oligodendrocyte transcription factor 2 (Olig2) and Neural/glial antigen 2 (NG2) antibodies were purchased from EMD Millipore (Billerica, MA). Myelin basic protein (MBP), neurofilament-Light chain (NFL), and GAPDH antibody were purchased from Cell Signaling Technology (Danvers, MA). Secondary anti-mouse-HRP and anti-rabbit-HRP antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX).

Immunoblots were performed as previously described [32] (link). Proteins were separated by SDS-PAGE (Bio-Rad), then probed with primary antibodies (rabbit anti-ADAR1 antibody from Zymed) followed by visualization with anti-rabbit HRP antibodies (Santa Cruz) and ECL plus (Amersham).