Blood urea nitrogen concentrations were measured with a Beckman Autoanalyzer (Beckman Coulter, Fullerton, CA). Serum anti-apoferritin IgG antibodies were measured by ELISA.9 (link) Briefly, 96-well plates were coated with apoferritin. Serial dilutions of sera were plated and incubated at room temperature for 2 hours, followed by HRP-conjugated goat anti-mouse IgG (Kierkegard & Perry Laboratories, Gaithersburg, MD) and OPD peroxidase substrate (Sigma-Aldrich, St. Louis, MO). The OD450 was then measured.
Circulating IC levels were measured by C1q binding ELISA as described previously.41 (link) Briefly, 96-well plates were coated with 10 μg/ml human C1q (Sigma) in carbonate buffer. After blocking with 1% BSA, sera samples were loaded in serial dilutions starting at 1/1000, followed by HRP-goat anti-mouse IgG (Sigma, 1:2000) and then TMB (Jackson ImmunoResearch Laboratories, West Grove, PA). The OD450 was then measured. Sera samples from unmanipulated wildtype mice were used as negative control.
Circulating IC levels were measured by C1q binding ELISA as described previously.41 (link) Briefly, 96-well plates were coated with 10 μg/ml human C1q (Sigma) in carbonate buffer. After blocking with 1% BSA, sera samples were loaded in serial dilutions starting at 1/1000, followed by HRP-goat anti-mouse IgG (Sigma, 1:2000) and then TMB (Jackson ImmunoResearch Laboratories, West Grove, PA). The OD450 was then measured. Sera samples from unmanipulated wildtype mice were used as negative control.