Dengue virus plaque development in simian cell systems. I. Factors influencing virus adsorption and variables in the agar overlay medium.

Chemical and physical variables influencing the plaquing of all dengue serotypes in two simian cell systems were studied. Calf serum in the nutrient overlay may be replaced by mouse ascitic fluid or bovine plasma albumin when employing the rhesus monkey kidney LLC-MK(2) cell system for plaquing all dengue serotypes. Doubling the serum concentration in the overlay had little effect in modifying dengue types 1, 2, 3, and 4 plaque titers. Newborn agamma, 4-week-old and 8-week-old calf serum gave comparable titers with all dengue virus serotypes. Dengue virus titers, plaque size, and development time were unaffected by sodium bicarbonate concentrations ranging from 1.1 to 4.4 mg/ml of overlay. A twofold increase (0.00332 g%) in the amount of either autoclaved or filtered-sterilized neutral red reduced the dengue 2 virus titer as much as 2.2 logs. An increased Mg(++) and decreased Ca(++) concentration in the overlay medium increased the efficiency of the plaquing system.