Study On Immune Detecting Technique Of Fish Lymphocystis Disease Virus

Lymphocystis disease is one of fish viral diseases, which is widely spreading and has made severe economic loss every year. In this experiment, immunofluorescence antibody technique ( IFAT ) and labelled streptavidin biotin ( LSAB ) immunohistochemical technique are used for locating lymphocystis disease virus (LCDV) in fish, and the way of virus infecting is also discussed.The virus as antigen is separated from lymphocystis nodules of flounder (Paralichthys olivaceus) surface and purified by discontinuous density gradient centrifugation method. By cell fusion technique, four positive hybridomas (2A9, 1A8,1B2,3G3 ) are raised and cloned which produce monoclonal antibodies (Mabs) against LCDV. These four Mabs can combine with LCDV of flounder specifically and not cross-react to tissues.Ten tissues of flounder and black rockfish (Sebastes fuscescens) infected by LCDV naturally are detected by IFAT and LSAB using Mabs. The results show that LCDV positive signals are found in stomach, intestine, gill and epidermis tissues of flounder with lymphocystis nodules on body surface, within which the signals are on the epithelium of stomach and intestine, around the second gill lamellae of gill and in subcutaneous muscle fibers of epidermis tissues; there are positive signals in stomach, intestine and epidermis tissues of flounder without obvious symptoms cultured in the same tank, and the locations of signal in tissues are same with that serious, but positive signals are not found in gill tissues; in black rockfish, positive signals are in stomach, gill and epidermis tissues and the locations of signal are similar with flounders; the results of detection in healthy flounder and black rockfish are negative. All these indicate that the two methods can be used to diagnose lymphocystis disease of founder efficiently, especially that without obvious symptoms. There is a possibility that LCDV enter fishes through digestive tracts and don't infect gill tissue at first but when the disease becomes more serious.Furthermore, the four monoclonal antibodies against LCDV of flounder are also used by IFAT to study lymphocystis disease black rockfish. It is turned out that three Mabs (2A