Complement-dependent cytotoxicity (CDC) of anti-FLRT2 antibody. CDC activities using two healthy control IgG and two anti-FLRT2 positive IgG, B11-8 and X10-48, at a concentration of 1.28 mg/ml with 1:3 diluted complement (A), 1.28 mg/ml of IgG, and 1:3 diluted complement with recombinant FLRT2 at the indicated concentrations (B), and various IgG concentrations with 1:6 diluted complement (C) against HUVECs were measured with the WST-1 assay. (D) CDC activities against mock transfected HEK293T cells (empty, left) and FLRT2-expressing HEK293T cells (FLRT2, right) by using 1.28 mg/ml of IgG and 1:3 diluted complement were measured with the WST-1 assay. HEK293T cells negative or positive for FLRT2 expression were stained with anti-FLRT2 antibody followed by secondary antibody against human IgG1, IgG2, IgG3, IgG4 (E), and IgM (F), and analyzed with flow cytometry. (G) ADCC activities using control IgG, B11-8, and X10-48, at a concentration of 1.28 mg/ml with an E:T ratio of 25:1 were determined with the lactate dehydrogenase detection method. Error bars indicate SD. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.
Shirai et al. Arthritis Research & Therapy 2012 14:R157 doi:10.1186/ar3897