Figure 2

Antibody responses of immunized sheep. (A) Immunofluorescence analysis of BTV-8 infected cells. Vero cells were infected with an MOI of 1 and further incubated for 24 h at 37 °C. The cells were fixed, permeabilized, and incubated with immune sera from sheep immunized twice with the indicated VRPs. The primary antibodies were detected with anti-sheep IgG-Alexa 546 conjugate (red fluorescence). Scale bar represents 24 μm. (B) Detection of BTV-8 neutralizing serum antibodies. Sera were prepared from VRP-immunized sheep (group size n = 6) at day 0 (first vaccination), day 21 (second vaccination), and day 42 (three weeks after the second immunization). Neutralization of virus was estimated 3 to 5 dpi according to the development of CPE. Neutralizing titres were calculated and expressed as ND₅₀/100 μL. Values equal to or lower than 2 were regarded negative.