Succinyllysine polyclonal rabbit antibody

The equal amounts of protein extracts (60 μg) were subjected to 12% SDS-PAGE and electrotransferred from SDS-gel to polyvinylidene difluoride membrane (GE Healthcare). After that, the membrane was blocked with 5% (w/v) bovine serum albumin in Tris-buffered saline with Tween-20 at ambient temperature for 2 h, and then succinyllysine polyclonal rabbit antibody (1:1000 dilution; PTM Biolabs, Inc) was added at 4 °C overnight, following the primary antibodies, washing the membrane with Tris-buffered saline with Tween-20 (25 mM Tris–HCl, pH 8.0, 125 mM NaCl, and 0.1% Tween-20), 15 min for three times. The membrane was incubated with horseradish peroxidase–conjugated anti-rabbit immunoglobulin G (1:2000 dilution) (Abgent) for 2 h at room temperature. Finally, the chemiluminescence of immune complexes was detected by using G:BOX Chemi XT4 system (Syngene).

The protein extracts (50 μg) were separated by 12% SDS/PAGE and transferred to a polyvinylidene difluoride (PVDF) membrane (GE Healthcare, Piscataway, NJ). The membrane was blocked at ambient temperature for 2 h in TBST (25 mm Tris·HCl, pH 8.0, 125 mm NaCl, 0.1% Tween 20) containing 5% (w/v) BSA. Then, the membrane was incubated with the succinyllysine polyclonal rabbit antibody (PTM Biolabs, Inc.) at a 1:2000 dilution overnight at 4 °C. After washing with TBST four times for 15 min each, the membrane was incubated with horseradish peroxidase-conjugated antirabbit IgG (1:2000 dilutions, ABGENT) at ambient temperature for 1 h. The G:BOX Chemi XT4 system (SYNGENE) was used for signal detection.