Optimization of suspended cell method and comparison with cell monolayer technique for virus assays.

The suspended cell technique for enumeration of viruses from environmental samples was evaluated in regards to the number of cells necessary per bottle or flask, contact time with virus prior to overlay, and number of viruses per sample for maximum enumeration, using 40-cm2 (6 oz) glass bottles or 25-cm2 plastic culture flasks. Optimum cell numbers were determined to be 4 x 10(7) cells for the 40-cm2 bottles and 2.0 x 10(7) cells for the 25-cm2 flasks. The optimum exposure time of virus to cells at 37 degrees C was 60 min with significantly higher recoveries with shaken mixtures as opposed to static mixtures. Upon comparing the suspended cell technique with that of the cell monolayer system, using two sets of environmental sewage sample concentrates, 62 of the 67 samples yielded an average of 5-8 times greater viral recoveries with the suspended cell procedure. Based on our data we feel that all environmental samples should be tested using the suspended cell procedure.