IN VITRO BINDING BETWEEN WHITE SPOT SYNDROME VIRUS AND HEMOCYTES OF PENAEID SHRIMP

White spot syndrome virus (WSSV-was isolated from the cephalothorax of infected Penaeus chinensis by differential centrifugation and then labeled by DIG-NHS in order to test whether it could bind to hemocytes of penaeid shrimp in vitro.Using an NBT/BCIP color detection system, dark purple grains were seen around cells by light microscopy and indicated stable binding between DIG-WSSV and hemocytes.Characterization of binding by holding the concentrat ion of DIG-WSSV constant and measuring absorption at 492nm with a perroxidase ( POD-detection system revealed that binding increased gradually as the quantity of hemocytes increased.Saturation was reached at 1.2 104 hemocytes per well.If the quantity of hemocytes was held constant, absorption ( binding-increased gradually as the quantity of DIG-WSSV increased and saturation was reached at 4g protein/well.Addition of sufficient unlabeled WSSV completely blocked this binding.Further analysis revealed that absorption at 4 ℃ increased with incubation time up to 40 min after which it decreased with prolonged incubation.Osmolarity of the binding buffer had little influence on binding while pH in the acid range promoted it.Incubation at 37℃ did not affect binding but at 55 and 70 ℃, it was reduced.Short term ultrasonic treatment increased binding but long term treatment reduced binding.Organic so-l vents reduced binding and this was particularly strong with chloroform/methyl alcohol.With respect to detergents, SDS and deoxycholic acid sodium salt decrease binding but Triton X-100 and Nonidet P-40 increased it.