Rotavirus concentration from cell culture harvests: trypsin treatment followed by hydroextraction

Abstract

An 80-to 150-ml amount of calf or simian rotavirus-containing cell culture harvests of MA-104 cells were treated with 50 jig of trypsin per ml and hydroextracted overnight (4°C) with polyethylene glycol 6,000. The concentrate was resuspended in 8 to 10 ml of tryptose phosphate broth and plaque assayed. Between 85 and 97% of the input virus could be recovered with a concentration of up to 15-fold. Earlier work in our laboratory showed that polyethylene glycol hydroextraction is a useful method for the second-step concentration of entero-(9), reo-(9), and rotaviruses (11) from the water environment. Because of the relatively simple and efficient nature of this technique, we attempted to extend its use to the concentration of rotaviruses from cell culture harvests (CCH). This communication describes the findings of these experiments. Trypsin (1:250; GIBCO, Grand Island, N.Y.) and soybean trypsin inhibitor (Sigma Chemical Co., St. Louis, Mo.) were prepared separately as