Rabbit anti ha

HA-tagged hM3Dq DREADD expression in the hippocampus and cortex of CamKIIα-tTA::TetO-hM3Dq bigenic mice (P7) was visualized using immunofluorescent staining for the HA epitope. Pups single-positive for either CamKIIα-tTA or TetO-hM3Dq were used as the genotype-controls. Mice were sacrificed by transcardial perfusion with 4 % paraformaldehyde, and 40 µm thick serial coronal sections were obtained using a vibratome (Leica, Germany).
Following a permeabilization step at room temperature in phosphate-buffered saline with 0.4% Triton X-100 (PBSTx) for one hour, the sections were then incubated in the blocking solution [1% Bovine Serum Albumin (Roche, Cat. No. 9048-49-1), 5% Normal Goat Serum (Thermoscientific, Cat. No. PI-31873) in 0.4% PBSTx] at room temperature for one hour. The sections were incubated with primary antibody, rabbit anti HA (1:250; Rockland, Cat. No. 600-401-384, USA) for 4 days at 4°C. Following sequential washes with 0.4% PBSTx, the sections were incubated with the secondary antibody, goat anti rabbit IgG conjugated to Alexa Fluor 568 (1:500; Invitrogen, Cat. No. A-11079, USA) for two and a half hours at room temperature, followed by washes with 0.4% PBSTx. Sections were mounted on to slides using Vectashield Antifade Mounting Medium with DAPI (Vector, H-1200, USA) and images were visualized on a LSM5 exciter confocal microscope (Zeiss, Germany).