Inactivation of recombinant baculovirus with binary ethylenimine (BEI).
Kidwaro, Fanson
Previous studies have shown the usefulness of BEI as an effective
inactivating agent for Baculoviruses. The goal of this study was to
develop a method for the complete and rapid inactivation of a
recombinant baculovirus (rBV) used in a veterinary vaccine. An aqueous
suspension of rBV was treated with 0 (control), 1.0, 2.5, 5.0, 7.5 and
10 mM BEI for 96 hours at 37[degrees]C. Aliquots were collected at
different times from 0 to 96 h and titrated on the Spodoptera frugiperda
cell line Sf9 to measure the inactivation kinetics. A Proof of
Inactivation Assay was developed using Sf9 cells to test for the
complete inactivation of rBV. Sf9 cells were grown in EX-CELL 420 insect
serum-free medium and inoculated 24 hours after planting with
BEI-inactivated harvest fluids. The inoculated cultures were incubated
for 120 hours at 27[degrees]C and examined for signs of cytopathic
effect (CPE) due to baculovirus. Cultures that was negative for virus
growth by CPE after initial inoculation and one blind passage were then
tested by PCR and Western blot. Our Proof of Inactivation Assay was able
to detect 3.0 infectious units after three passages. We have
demonstrated that 10 mM BEI was capable of inactivating 6.0 TCID50 per
ml of rBV within 24 hours. The results of this study led to the
optimization of BEI inactivation of rBV for use in a veterinary vaccine.
* Lenz, M.C., F.D. Worman, C. Bracken, R.A. Hesse and K.M. Mulkey.
Department of Agriculture, Central Missouri State University.
