Figure 4.

Heterologous and specific IgM binding of sera from cyprinid herpes virus 3 (CyHV-3) infection-survivor carp populations to consecutive frg11_VHSV and frgII_CyHV-3. Individual serum was obtained from CyHV-3 infection-survivors and healthy carp populations, diluted 100-fold and assayed in the same plate by consecutive frg11_VHSV + frgII_CyHV-3 enzyme-linked immunosorbent assay (scheme in Figure 3). Two micrograms of recombinant frg11_VHSV or frgII_CyHV-3 were used as solid phases. Heterologous and specific IgM binding of sera from carp surviving infection with CyHV-3 was estimated after preincubation of the sera with E. coli extract at an optimal concentration to lower to ~0.2 absorbance units the mean IgM binding of sera from the healthy carp population. Each raw absorbance value was normalized by the formula, raw individual absorbance value × 0.2/mean of healthy sera (n = 65). The IgM binding assays were repeated two to three times and their individual means used for further calculations. After normalization, the values were distributed in 0.08 absorbance classes and polynomically fitted, as indicated in Section “Materials and Methods.” *Statistically significant differences at the p < 0.05 level (Student’s t-test). (A) Heterologous and specific IgM binding of sera from the CyHV-3 infection-survivor carp population. Dotted black line (gray surface)—heterologous IgM binding of sera from the CyHV-3 infection-survivor carp population to frg11_VHSV (n = 91). Black line—specific IgM binding of sera from the CyHV-3 infection-survivor carp population to frgII_CyHV-3 (n = 91). Black dotted and dashed lines—heterologous and specific IgM binding from the healthy carp sera population, respectively, normalized to 0.2 absorbance means (n = 65). (B) Lack of correlation between heterologous (X-axis) and specific (Y-axis) IgM-binding concentrations from the individual sera from the CyHV-3 infection-survivor carp.