Anticardiolipin elisa

Autoreactivity was determined by ANA Hep-2 Staining Analysis (ZEUS Scientific Cat. No: FA2400) and anticardiolipin ELISA (Inova Diagnostics Cat. No.: 708625). All antibodies were tested at 25 and 50 μg/ml as per manufacturer’s protocols. Scores from 0 to 3 were defined with four control antibodies VRC01-LS, 4E10, VRC07-523-LS, and VRC07-G54W. Hep-2 cells (human malignant epithelial cells, female, obtained from ZEUS Scientific). Signals of the test antibody were visually estimated in comparison to the control ones. Scores greater than 1 at 25 μg/ml were classified as autoreactive, and between 0 and 1 as mildly autoreactive. In the cardiolipin ELISA, bNAbs were tested at a starting concentration of 100 μg/ml, followed by 3-fold dilutions. IgG phospholipid (GPL) units were calculated from the standard curve. GPL score < 20 was considered as not reactive, 20–80 as low positive and >80 as high positive.

Autoreactivity was determined by ANA Hep-2 Staining Analysis (ZEUS Scientific Cat. No: FA2400) and anticardiolipin ELISA (Inova Diagnostics Cat. No.: 708625). For the Hep-2 assay, all antibodies were tested at 25 and 50 μg/ml as per manufacturer’s protocol and imaged on a Nikon Ts2R microscope for 500 ms. Scores from 0 to 3 were defined with four control antibodies VRC01-LS, 4E10, VRC07-523LS, and VRC07-G54W. Test antibodies were scored by visual estimation of staining intensity in comparison to the control antibodies. Scores equal to or greater than 1 at 25 μg/ml were classified as autoreactive, and between 0 and 1 as mildly autoreactive. In the cardiolipin ELISA, antibodies were tested at a starting concentration of 100 μg/ml, followed by threefold dilutions. IgG phospholipid (GPL) units were calculated from the standard curve. GPL score < 20 was considered as not reactive, 20–80 as low positive and > 80 as high positive.