Hrp conjugated donkey anti rabbit igg antibody

Neuro2a cells were lysed in radioimmunoprecipitation assay buffer (Nacalai Tesque). SDS/polyacrylamide gel electrophoresis (PAGE) was performed using 8% and 10% SDS/polyacrylamide gel to detect CPEB1 and glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH), respectively. In the case of detecting CPEB1, SDS/PAGE was performed until a prestained molecular marker protein (50 or 58 kilodalton) was fully run to the bottom of the gel to separate CPEB1 from adjacent nonspecific bands. Subsequent western blot analysis was performed using Trans‐Blot Turbo Transfer System (Bio‐Rad Laboratories, Hercules, CA, USA). Primary antibodies and horseradish peroxidase (HRP)‐conjugated secondary antibodies were used at dilutions of 1 : 500 and 1 : 1000, respectively, in 5% Blocking One (Nacalai Tesque) in Tris‐buffered saline containing 0.1% Tween 20. Proteins were visualized using Luminata Classico Western HRP Substrate (Millipore, Burlington, MA, USA) and detected using ImageQuant LAS 4000 mini (GE Healthcare). The primary antibodies used in this study were rabbit polyclonal antibodies against CPEB1 (ab73287, Abcam, Cambridge, UK) and GAPDH (sc‐25778, Santa Cruz Biotechnology, Santa Cruz, CA, USA). The secondary antibody was HRP‐conjugated donkey anti‐rabbit IgG antibody (Jackson ImmunoResearch, West Grove, PA, USA). The raw images of western blots are shown in Fig. S1 and Fig. S2.