Serum anti-dsDNA antibodies were detected by enzyme-linked immunosorbent assay (ELISA) as previously described (15 (link)). Briefly, 96-well plates were precoated with methylated bovine serum albumin (BSA) at a concentration of 10 mg/ml, followed by 5 μg/ml of calf thymus dsDNA (Sigma-Aldrich). After blockade with 1% BSA, sera were added in serial dilutions and incubated for 1 hour at room temperature. After washing, horseradish peroxidase–conjugated goat anti-mouse IgG antibody (Sigma-Aldrich) was added to detect the bound anti-dsDNA, followed by the peroxidase substrate tetramethylbenzidine. The reaction was terminated with 1M H2SO4, and the absorbance at an optical density of 450 nm was determined. Normal mouse IgG was used as a negative control.
Levels of IL-1β, IL-17, and IFNγ were determined with ELISA kits (R&D Systems) according to the manufacturer's instructions.
Levels of IL-1β, IL-17, and IFNγ were determined with ELISA kits (R&D Systems) according to the manufacturer's instructions.
