Identification of mimotope peptides which bind to the SARS-CoV spike protein specific monoclonal antibody 2C5 with phage-displayed peptides library / 生物工程学报

To identify the epitope of SARS-CoV spike protein specific neutralizing monoclonal antibody (MAb) 2C5. The antibody was used as target and three rounds of bio-panning were conducted with phage-display peptide library. After the third panning, 20 phage-plague clones were randomly picked and analyzed for the binding ability with the MAb 2C5 by ELISA. The display sequence analysis demonstrated that among the twenty phage clones, eight clones displayed the same seven-peptide TPEQQFT. All these eight phage-clones showed strongest binding activity with 2C5 in phage ELISA analysis. Furthermore, phages displaying peptide TPEQQFT could specifically inhibit the binding of MAb 2C5 with SARS-CoV spike protein. The results demonstrated that TPEQQFT is a mimic epitope peptide containing neutralizing MAb 2C5. This study may provide information for further structural and functional analysis of spike protein and development vaccine for severe acute respiratory syndrome.

Expression of truncated gE gene of pseudorabies virus (PRV) and primary application in differential diagnosis of PRV vaccination and infection / 生物工程学报

With the application of gE gene deleted pseudarabies virus (PRV) vaccine worldwide, a corresponding differential diagnosis based on gE glycoprotein is needed in the project of PRV eradication. In this study, PRV gE gene without signal and transmembrane region was amplified by PCR and cloned into pGEX-6P-1, generated pGEX-gE. After transformation of BL21 with pGEX-gE, an expressed fusion protein(about 63kD) was identified by SDS-PAGE. The recombinant proteins are produced as inclusion bodies. By changing the inductive conditions, the formation of inclusion bodies was inhibited and tended to increase the percentage of soluble recombinant protein. The antigenic reactivity of the recombinant protein was confirmed by Western blotting with polyclonal antibodies against PRV. Using purified recombinant tgE as antigen, an ELISA was developed to detect sera of PRV infected pigs and sera of pigs immunized with gE-deleted PRV vaccine. The total of 400 serum samples collected from field were comparatively tested with the tgE-ELISA and a commercial competitive ELISA based on monoclonal antibody against gE, the results indicated that the coincidental rate between the two tests is about 94%.