Rabbit anti mouse insulin antibodies

Immunofluorescence staining was carried out after acetone fixation, permeabilization, and blocking with Avidin/Protein blocking together with 5% goat serum using rabbit-anti-mouse insulin antibodies (Cell Signaling, 1:100) and donkey-anti-rabbit^{AlexaFluor647} antibodies (Dianova, 1:400). For CD3 staining, arm.hamster anti-mouse antibodies (BD, 1:50) were used, followed by goat-anti-arm.hamster antibodies conjugated with Dylight⁴⁸⁸ (Dianova, 1:100). For Tet2 staining, rabbit-anti-Tet2 antibodies (ABclonal, clone A5682, 1:25) was used with biotinylated horse-anti-rabbit antibodies (Vector, 1:100) combined with SA^{Dylight 549}. For Foxp3 staining, cells were incubated with rat-anti-mouse Foxp3 antibodies (eBioscience, clone FJK-16s, 1:50) and biotinylated goat anti-rabbit (BD), combined with SA^{Dylight 549}. Nuclei were counterstained with DAPI (Diavona). For Tgfbr1 staining, rat-anti-mouse/human Tgfbr1 antibodies (R&D, clone 141231, 1:25) was used, followed with biotinylated goat-anti-rat antibodies (1:250) combined with SA^{Dylight 549} (Dianova, 1:200). Negative control slides were incubated with secondary antibodies. Cells were analyzed by confocal microscopy (Zeiss LSM700).