Fitc gl7 ra3 6b2

Mouse spleens or kidneys were embedded in OCT compound and snap frozen over liquid nitrogen. 5μm thin sections were cut on a cryostat, mounted on ColorFrost plus microscope slides (Fisher Scientific, PA) and fixed in cold acetone for 20 min. The following antibodies and reagents were utilized for immunofluorescence staining of mouse spleen sections for germinal centers: PE-anti-CD4 (GK1.5, Biolegend); FITC-GL7 (RA3–6B2, BD Biosciences); APC-anti-IgD (11–26c2a, BD Biosciences). Kidney sections were stained for C3 using FITC-anti-C3 (ICL Laboratories) or Biotin-anti-IgG (Jackson Immunoresearch) followed by SA-PE. Anti-nuclear Ab (ANA) reactivity was detected by indirect immunofluorescence staining of HEp-2 cell slides using sera from indicated mice at a 1:50 dilution, and probed with FITC-rat anti-mouse Kappa (H139–52.1). The images of stained spleen and kidney sections were captured using the Leica DM4000 fluorescence microscope and analyzed using a Leica application suite-advanced fluorescence software (LAS-AF, Leica Microsystems). For the measurement of the GC area, a total of at least 10 randomly selected germinal centers (GL-7⁺) per mouse spleen section from at least 5 mice of each genotype were measured for total area (μm²) using the LAS-AF quantitation tool.

Mouse spleens or kidneys were embedded in OCT compound and snap frozen over liquid nitrogen. 5μm thin sections were cut on a cryostat, mounted on ColorFrost plus microscope slides (Fisher Scientific, PA) and fixed in cold acetone for 20 min. The following antibodies and reagents were utilized for immunofluorescence staining of mouse spleen sections for germinal centers: PE-anti-CD4 (GK1.5, Biolegend); FITC-GL7 (RA3-6B2, BD Biosciences); APC-anti-IgD (11-26c2a, BD Biosciences). Kidney sections were stained for C3 using FITC-anti-C3 (ICL Laboratories) or Biotin-anti-IgG (Jackson Immunoresearch) followed by SA-PE. Anti-nuclear Ab (ANA) reactivity was detected by indirect immunofluorescence staining of HEp-2 cell slides using sera from indicated mice at a 1:50 dilution, and probed with FITC-rat anti-mouse Kappa (H139-52.1). The images of stained spleen and kidney sections were captured using the Leica DM4000 fluorescence microscope and analyzed using a Leica application suite-advanced fluorescence software (LAS-AF, Leica Microsystems). For the measurement of the GC area randomly selected germinal centers (GL-7⁺) were measured for total area (μm²) using the LAS-AF quantitation tool.