Western blot reactivity patterns.

*Blot analysis criteria for human diagnostics provided by the manufacturer (Microgen, Neuried, Germany): A positive band was assigned the following point values: NS3, 3; Helicase, 3; NS5-12, 2; NS4, 4; Core, 8. Only weakly positive bands (visible, but weaker than the core antigen of the weak positive HCV control) were considered negative with the exception of the core antigen, which was then assigned 5 points. Blots were considered positive if the sum of points was equal to or greater than 10, borderline if the sum was between 6 and 9 points, and negative if the sum was equal to or below 5 points. Eight bat sera showing strongest Western Blot reactivity were end-point diluted to evaluate specificity of the reaction. Bands remained visible up to a dilution of 1∶400. Some analyses were done using a follow-up version of the Microgen assay (recomline) with two separate core antigens and different interpretation criteria. Blots were then considered borderline if any two antigens showed higher intensity than a cut-off control, the helicase alone, the helicase and any NS antigen or any core epitope. Blots were considered positive in this assay version if both core antigens were positive, one core plus any other antigen or if three antigens showed higher intensities than the cut-off control. Reactivity of the secondary goat anti-mouse and tertiary donkey anti-goat antibodies used for rodent testing were controlled by using mouse anti-core and anti-NS3 monoclonal antibodies for primary reaction with blot antigens.