Establishment Of Combined-antigen ELISA And Epidemiological Study For Human Herpesvirus 8 Infection In Xinjiang

Objective: To construct prokaryotic expression plasmids of pET_41a-ORF_73CC, pET_41a-ORF₆₅, pET_41a-K8.1, and to purify and identify their fusion proteins. The three fusion proteins were used to establish a combined-antigen ELISA assay for Kaposi's Sarcoma-Associated Herpesvirus (KSHV) seroepidemical survey in Xinjiang. In addition, initial analysis for understanding the current epidemic situation and risk factors were made in relation to HHV-8 prevalence in Xinjiang. Methods: Using recombined plasmids pGEM-Teasy/ORp73 , pGEM-Teasy/ORF₆₅, pGEM-Teasy/ORF_K8.1 as the templates, EcoR I and Xho I restriction digestion sites were amplified by PCR and subsequently inserted into prokaryotic vector pET-(41a) (+). And then identified by double enzyme digesting and sequencing. The recombinant plasmids pET_41a-ORF_73cc, pET_41a-ORF₆₅, pET_41a-K8.1 were trans-formed into expression vector E.coli BL₂₁(DE3). The fusion proteins were expressed by induction with IPTG. The expressed recombinant proteins were purified using poly-adenylic histiding affinity chromatography. Purification of the target proteins were confirmed by SDS-PAGE and Western-Blot. The expressed recombinant proteins were used as antigens constructing an enzyme-linked immunosorbent assay (com-bined-antigen-ELISA), The seram from 44 KS patients and 23 skin cancer were used to identify the sensitivity and specificity of the combined-antigen-ELISA assay; randomly questioned tumur patients in First Teaching Hospital and Tumur...