Construction And Application Of The Expressive Clone Of Human Herpesvirus 7 Pp85

Objective To construct prokaryotic expression clone of human herpesvirus 7 (HHV7) pp85, and to prepare of efficient recombinant antigen for serological diagnosis of HHV7 active infection.Methods ①SupTl cells were infected with HHV7 (Glasgow ) and harvested. ② The major antigen epitopes coding sequence( 328 ~ 533 aa) of HHV7 ORF U14 gene was amplified by PCR and inserted into the prokaryotic expression vector pThioHis A, recombinant plasmid was transformed into E.coli BL21 and induced to express by IPTG. The products were purified by nickel-chelating Sepharose resin affinity chromatography. The antigenicity and the specificity of the recombinant proteins was identified by Western blot. ③ ELISA coating with the cells lysates and the recombinant proteins was used to screen 630 serum samples, and 325 of them simultaneously detected by ELISA kit (Qiagen). The specificity, sensitivity, Youde-n's index, predictive value positive and predictive value negative of the cells lysates and the recombinant proteins were detected. The cross-reactivity with other β human herpes-virus were also detected.Result ELISA coating with the cells lysates and the recombinant proteins had a good agreement with ELISA kit (Qiagen), and had no cross-reactivity with other β human herpesvirus . The ELISA coating with the cells lysates and the recombinant proteins showed high specificity and sensitivity in the detection of HHV7-specific IgG and IgM antibodies in clinical samples. The recombinant protein was more specific and lower sensitive than the cells lysates.Conclusion The cells lysates and the recombinant proteins can be used in the serological diagnosis for acute and active HHV7 infection. Recombinant prokaryotic expression vector was gained, and it is valuable for advanced study on HHV7 specific antibody detection.