| Severe acute respiratory syndrome is a new infectious disease, which hasimpacted on human being's normal life and stability of the society. According to thedata, there were 8437 persons being infected over the world and in which 813 patientsdied from the disease until July 11, 2003. The overall mortality rate of the disease wasabout 10.5%. Since the infectious disease of SARS spread by the respiratory route anduntil now there are no effective antiviral drugs and vaccines to treat the disease. Thenit is very important to have a rapid and accurately diagnostic tool to detect SARS-CoV at early stages of the disease.We expressed and purified full length and different truncated fragments of SARSnucleocapsid( N ) proteins. Rabbits and BALB/c mice were immunized the full-length of N protein to induce SARS N protein-specific polyclonal antibody andmonoclonal antibody. The monoclonal antibodies specifically recognized both ofE.coli expressed N protein and native N protein from the lysates by Western Blotting,which suggested that the mAbs were able to recognize native N protein of SARS-CoV.To locate the epitopes of giving Abs, five recombinant N protein fragments wereselected and their binding activities to raise six mAbs were tested by ELISA. Theresults showed that all of raised mAbs ( six clones ) were divided into two groups,which recognized the regions of amino acids 249-317 ( A group ) or 317-395 ( Bgroup ). The recognizing region spanning 249-395 is the predominant B cell epitopeslocated at the C-terminus of N protein. One pair-Abs, N protein-specific polyclonalantibody and mAb, were selected to construct a sandwich ELISA-kit. The kit was ableto detect N protein with a liner curve. It was also to observe that all of selected patientsera, including tumor, systemic lupus erythematosus( SLE ), syphilis, hepatitis Bvirus( HBV ) and hepatitis C virus( HCV ), did not react with the kit. The experimentssuggested that this kit had a potential in detecting N protein of SARS-CoV. |
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