Fig. 4. ELISA of OspC variants with variant-specific sera.

Fifteen sera (“sA” through “sU”) from mice, each immunized with a specific recombinant OspC variant, were previously assayed for reactivity with the 16 OspC variants (“rA” through “rU”) using ELISA [48]. (Top) Each panel shows binding intensities (normalized z-scores, y-axis) of an OspC variant with a panel of OspC-specific sera (x-axis). Error bars show one standard deviation above and below the mean from two replicated assays. A value above the z = 2 line (dashes) indicates a highly significant reaction. (Bottom left) A heat map representation of the mean z-scores. (Bottom right) Antigen reaction characteristics (ARC) curves, similar to the receiver-operation characteristics (ROC) curves, is a measure of antigen specificity. Each curve traced the cumulative z-scores (y-axis) of an OspC variant’s binding intensities with the sera samples, ordered by the lowest to the highest reactivity. The ARC curve rises with an above-average binding value (z > 0) and drops with a below-average binding value (z < 0). Thus, a high-rising curve (e.g., for rB) indicated consistently above-average reactivity with sera samples, suggesting a broadly cross-reactive antigen. Conversely, a low-lying curve (e.g., for rG) indicated consistently below-average reactivity, suggesting a relatively specific antigen. Curves close to the zero line (the majority of variants) indicated antigens with an average level of cross-reactivity.