Goat anti mouse igg

CD4⁺ T cells, antigen-presenting cells, Treg cells and neutrophils were each purified from the splenocytes of 4-week-old BDC2.5⁺Il-10^+/+ NOD mice, BDC2.5⁺Il-10^-/- NOD mice, or wild type NOD mice. Splenic CD4⁺ T cells were purified by depletion, of CD8⁺ T cells (clone T1B105), MHC class II⁺ cells (clone 10.2.16), and B cells (anti-mouse IgM and IgG), incubating the cells with monoclonal antibody (mAb) hybridoma supernatants, followed by magnetic bead separation. For splenic antigen-presenting cell (APC) isolation, anti-Thy1 (Y19) mAb hybridoma supernatant and complement was used to remove Thy1⁺ T cells. The supernatants of different mAb hybridomas were kindly provided by the late Charles Janeway (Yale University). Magnetic beads conjugated with goat anti-mouse IgG, goat anti-mouse IgM, or goat anti-rat IgG were purchased from QIAGEN. Treg cells were isolated using MojoSort™ Mouse CD4⁺ T cell Isolation Kit (BioLegend), followed by CD25 positive isolation using a PE-anti-mouse CD25 antibody (Clone, PC61, BioLegend) and MojoSort™ Mouse anti-PE Nanobeads (BioLegend). The remaining CD4⁺CD25^- T cells were used as effector CD4⁺ T cells. Neutrophils were isolated according to the manufacturer’s instructions using MojoSort™ Mouse Neutrophil Isolation kit (BioLegend).