Preparation And Preliminary Application Of Fluorescent Antibody To H5Nl Avian Influenza Virus

Avian influenza is a group of symptoms caused by influenza virus A.Avian influenza has raised great.concerns for its enconomic importance to poultry industry and potential threat to the public health.Recently,the widespread nature of H5N1 arise highly attention.So a accurate,rapid and hypersensitive technique is in concerned. Monoclonal antibody against avian influenza virus hemagglutination was developed by means of the conventional protocol.The IgG was purified by affinity chromatography and conjugated with FITC.Virus titer and distribution in infected mouse was detected by direct fluorescent antibody test(dFAT).The 8-week-old female Balb/c mice were immunized with the AIV HAB/01, which was purified and concentrated.The spleen cells of immunized mice were fused with SP2/0 myeloma cells.Indirect enzyme linked immunosorbent assay(ELISA) was used to screen hybridoma cells and limited dilution method was performed to subclone positive clones.Three of McAbs specifically against avian influenza virus (AIV) were obtained,designated as1B6,2A3 and 3A5 respectively.One of these 3 McAbs,1B6 was of IgM subtype.The others were of IgG1 subtype.The titers of ascites were:1:2~6,1:2~6,1:2~8.The results of the cross-reaction test suggest that these McAbs were specific to AIV-H5N1 without detectable cross-reactivity with NDV and other types avian influenza virus.Western-blot indicated the 3A5 is against HA protein.The McAb were purified by protein G and labelled with FITC.The results of absorption test and blocking test showed that the fluorescent antibody(FA) has highly specificity and sensitivity.It can use for AIV laboratory detection.