Goat anti rat igg alexa fluor 488

Antibodies for immunofluorescence were purchased from the following sources: F4/80 (0.2 μg/ml, BM8, OriGene, Rockville, MD, USA), TNF-α (2.5 μg/ml, MP6-XT3, eBioscience, San Diego, CA, USA), IL-10 (2.5 μg/ml, JES5-2A5, eBioscience), fibronectin (0.7 μg/ml, Polyclonal, invitrogen, Carlsbad, CA, USA), and collagen IV (2.5 μg/ml, Polyclonal, Invitrogen). Secondary antibody were goat anti-rat IgG Alexa Fluor® 488 (2 μg/ml, BioLegend, San Diego, CA, USA), streptavidin Alexa Fluor® 594 (0.5 μg/ml, BioLegend), goat anti-rabbit IgG Alexa Fluor® 488 (2 μg/ml, Invitrogen).
The following antibodies (BioLegend) were used for flow cytometry: APC anti-mouse F4/80 (2.5 μg/ml, BM8), PE/Cy7 anti-mouse CD86 (1.25 μg/ml, GL-1), PE anti-mouse CD206 (1.25 μg/ml, C068C2), PE/Cy7 anti-mouse TNF-α (5 μg/ml, MP6-XT2), and Alexa Fluor® 488 anti-mouse IL-10 (12.5 μg/ml, JES5-16E3).

For detection of antibodies and antibody/complement complex deposition in cardiac and kidney tissue in vivo, 5 µm sections of hearts and kidneys of Resiquimod-treated mice were stained with goat anti-mouse IgG-FITC (Sigma, F5387, lot 99H4872, 1:200), anti-mouse IgM-Alexa Fluor 488 (BioLegend, 406521, lot B203057, 1:200), or unlabelled anti-mouse IgG1/IgG2a/IgG2b/IgG3 (BioLegend, 406601/407101/406701/406802, 1:200) and a secondary goat anti-rat IgG-Alexa Fluor 488 (BioLegend, 405418, 1:500). C3 staining was performed using anti-mouse C3c-FITC (Thermo Fisher Scientific, PAI-29718, lot QL2122721H, 1:50), as described previously (Tarzi et al., 2002 (link)). Sections were counterstained with wheat germ agglutinin (WGA)-Alexa Fluor 594 (Thermo Fisher Scientific) to label membranes for 15 min at room temperature and mounted in Vectashield mounting liquid containing DAPI (Vector Laboratories, Peterborough, UK) to label nuclei.
Images were captured using a LMD7000 microscope (Leica microsystems) and processed using ImageJ.