Goat anti mouse texas red conjugated igg

LC3B, iNOS, and CD206 in KCs were identified by indirect immunofluorescence staining using rabbit anti-mouse LC3B mAb, anti-mouse iNOS mAb, and anti-mouse CD206 mAb (Cell Signaling Technology, MA, USA). After washing, the KCs which were premounted with VECTASHIELD medium with DAPI (Vector) were incubated with secondary antibody goat anti-mouse Texas Red-conjugated IgG (Sigma, Saint Louis, MO, USA). The numbers of positive-stained cells were counted blindly in 10 high-power field (HPF) sections.

LC3B, iNOS, and CD206 in KCs were identified by immunofluorescence using rabbit anti-mouse LC3B mAb, anti-mouse iNOS mAb, and anti-mouse CD206 mAb (Cell Signaling Technology, MA, USA). After incubation with secondary goat anti-mouse Texas Red-conjugated IgG (Sigma, Saint Louis, MO, USA), the KCs were pre-mounted with VECTASHIELD medium with DAPI (Vector). Positive cells were counted blindly in 10 HPF/section (200×). The positive cells were expressed as a percentage of total cells.

Tissues fixed with 4% formalin were embedded in paraffin. Then, tissue sections were incubated with primary antibody of SIRT3 (Invitrogen, Carlsbad, CA, USA) or FOXO3 (Proteintech, Wuhan, China). Biotinylated goat antirat IgG (Vector, Burlingame, CA, USA), as the secondary antibody, was incubated with immunoperoxidase (ABC Kit, Vector) following the manufacturer’s protocols. SIRT3 and FOXO3 in BMDMs were detected by immunofluorescence with antirabbit SIRT3 pAb (Invitrogen, Carlsbad, CA, USA) and antimouse FOXO3 mAb (Proteintech, Wuhan, China), followed by incubation with secondary goat antirabbit Texas Green-conjugated IgG or goat antimouse Texas Red-conjugated IgG (Sigma, St. Louis, MO, USA). DAPI was used to stain the nuclei. The slides were washed twice with PBS and detected using confocal microscopy (ZEISS, Oberkochen, Germany) in accordance to the manufacturer’s protocols. Positive cells were observed and counted in 10 HPF/section (×200 or ×400).