Goat anti mouse igm alexa 594

Mice were sacrificed and subjected to transcardial perfusion of phosphate-buffered saline (PBS) for 5 min at room temperature, followed by 4% paraformaldehyde in PBS for 15 min at 4°C. After perfusion, the lumbar DRGs were immediately removed and postfixed in 4% paraformaldehyde in PBS at 4°C overnight.
After postfixation, the samples were transferred to PBS containing 30% sucrose and dehydrated for 3 days. The tissues were then embedded in optimal cutting temperature compound and cut into 20-μm longitudinal sections using a cryostat. After treatment with 0.2% Triton X-100 for 5 min, the DRG sections were blocked with 5% normal goat serum (NGS; Vector Laboratories, Burlingame, CA) and then incubated with a mouse monoclonal anti-V1a receptor (1:300; LSBio, Seattle, US) and rabbit polyclonal anti-calcitonin gene-related peptide (CGRP, 1:300; BMA Biomedicals, Augst, Switzerland) or rabbit polyclonal anti-S100 (1:300, Leica Biosystems, Wetzlar, Germany) for 24 h at 4°C. Following incubation, the antibody was visualized with goat anti-mouse IgM Alexa 594 (1:300; Invitrogen, Carlsbad, CA) for 1 h at room temperature. Images were obtained on an AX70 Olympus microscope (Olympus Optical Co., Ltd., Tokyo, Japan). ImageJ (https://imagej.nih.gov/ij/, Wayne Rasband, NIH) was used to quantify the V1a-positive area in each model.