An agar overlay plaque assay method for autographa californica nuclear-polyhedrosis virus

Abstract

A sensitive and reproducible infectivity assay of the Autographa californica nuclear-polyhedrosis virus (Ac-NPV) using Trichoplusia ni (TN-368) tissue culture cells was developed. A mixture of Ac-NPV infectious tissue culture supernatant and cells was centrifuged at 1000g for 60 min. The supernatant was removed, and the cells were overlaid with medium containing 1.5% agarose. Virus polyhedral inclusion bodies (IB) were observed in cells after 24 hr. Plaques (areas of IB containing cells) measured 2-3 mm after 5 days postinoculation. Plaque numbers were directly related to inoculum concentration. The assay was found to have a higher sensitivity than standard TCID50 or methylcellulose overlay plaque assay procedures. © 1977 Academic Press, Inc.