Polyclonal rabbit anti human c3c

Complement C3 binding assay for bacterial cells was carried out by indirect ELISA according to Alberti et al. [12 (link)]. Complement C3 activation assay for LPS was performed by a direct sandwich ELISA according to Holmskov-Nielsen [13 (link)]. The assays were performed on the pool of HS with the correct of parameters of C3 complement protein and lacking specific anti-Salmonella antibodies. Polyclonal rabbit anti-human C3c (Dako) (2.5 μg/ml), and polyclonal goat anti-rabbit immunoglobulins/HRP (horseradish peroxidase-conjugated, Dako) were used to detect C3 activated fragments. The plates were developed with O-phenylenediamine dihydrochloride (SIGMAFAST™ OPD, Sigma-Aldrich) and measured at A₄₉₂.

Female BALB/C mice were obtained from Beijing Vital River Laboratory Animal Technology Co., Ltd. In the experiments outlined below, avidin from egg white (Aladdin), C1q (EMD chemicals), alkaline phosphatase (AP) substrate P-nitrophenyl phosphate (Sigma-Aldrich), Mouse total IgG (Sigma-Aldrich), and human total IgG (Sigma-Aldrich) were used. Furthermore, the following antibodies were adopted: AP-conjugated polyclonal goat anti-human IgG (γ-chain specific) (Sigma Aldrich), AP-conjugated polyclonal goat anti-mouse IgG (whole molecule, Sigma-Aldrich), AP-conjugated polyclonal goat anti-rabbit IgG (whole molecule, Sigma-Aldrich), and polyclonal rabbit anti-human C3c (Dako). C1q CLR was produced by partial pepsin digestion of C1q as previously described (13 (link)).