Anti mouse or anti rabbit igg antibody labeled with horseradish peroxidase

Proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using 10% or 12% acrylamide gels that were subsequently subjected to western blotting. After SDS-PAGE, proteins were blotted onto a polyvinylidene fluoride (PVDF) membrane using the Mini Trans-Blot Electrophoretic Transfer Cell (Bio-Rad, Hercules, CA, USA). After blocking in 5% skim milk with Tris-buffered saline containing 0.1% Tween 20 (TBST), the membrane was probed with 1:5000 rabbit anti-BmNPV GP64 polyclonal antibody (Biogate, Gifu, Japan). This antibody can recognize both AcGP64 and BmGP64. The membrane was washed with TBST and incubated for 1 h in 1:20,000 anti-mouse or anti-rabbit IgG antibody labeled with horseradish peroxidase (GE Healthcare, Buckinghamshire, UK). Detection was performed using ECL Plus Western blotting reagent (GE Healthcare Japan, Tokyo, Japan). Specific bands were detected on a Fluor-S MAX MultiImager (Bio-Rad). To detect GFP-specific green fluorescent protein bands, the samples were mixed with sample buffer21 (link) without boiling and separated by SDS-PAGE. Green fluorescent bands were detected using a Molecular Imager FX (Bio-Rad).