Standardization and kinetics of in vitro bovine blood lymphocyte stimulation with bovine rotavirus

Two groups of 3-month old calves were immunized intramuscularly with attenuated bovine rotavirus and boosted 21 and 42 days later. The first group of three calves were vaccinated with live virus emulsified with incomplete Freund's adjuvant (IFA) and the second group was immunized with live virus suspended in phosphate buffered saline (PBS). Three other calves, serving as controls, were inoculated with PBS emulsified with IFA. The specific cell-mediated and antibody responses of the animals were studied. Preliminary analysis of in vitro peripheral blood lymphocyte transformation to bovine rotavirus determined optimal conditions as: 96 h culture period, 5 X 10(5) cells per culture in RPMI 1640 medium containing 10% heat-inactivated bovine fetal serum and the use of inactivated virus in the cell culture at a concentration of 5 X 10(6) median tissue culture infective dose before inactivation. Specific blastic stimulation was observed on calves immunized with the rotavirus emulsified with IFA after the second and third vaccine inoculation with stimulation index values varying from 2.00 to 5.73. Serum neutralizing antibody titers of 1/25,600 were also induced in the same calves. Calves immunized with rotavirus-PBS suspension developed a mean antibody titer of 1/1,600, but showed no specific lymphocyte stimulation. No increase in specific immune responses was detected in the control animals.