A novel antigen immunochromatography fluorometric strip for rapid detection and application of pathogenic bacterial high-quality antibody.

Unlike traditional immunoassay strips, a novel antigen immunechromatography fluorometric strip (AICFS) using inactivated bacterial antigen instead of an antibody as a test line and goat anti-mouse IgG-FITC as a tracer was developed. The applicability survey of AICFS indicated that E. coli O157:H7 (D3) and Acidovorax citrulli (6F) hybridoma cell cultures could be detected, but Vibrio parahemolyticus (H7, C9) hybridoma cell cultures were missed compared with the indirect enzyme-linked immunosorbent assay (ELISA). The four antibody affinity constants (Ka) were measured and compared, and AICFS could be suitable for high-affinity antibody detection. Compared with the traditional indirect ELISA, the AICFS sensitivity for D3 cell cultures, ascites, and purified antibodies was at least 2-fold more sensitive, the AICFS specific for D3 cell cultures by comparative interpretation was compliant except for the strain ATCC 43895, and the indirect ELISA missed it. More importantly, the AICFS method was confirmed by various real samples that it could be used in different scenarios regarding the antibody, including McAb preparation, the effective antibody use, and high-affinity antibody-secreted hybridoma auxiliary preparation and screening. It could be an excellent alternative method with less than 5% corresponding processing time for indirect ELISA method for pathogenic bacterial high-quality antibody detection. This is the first report of using AICFS for bacterial high-quality antibody detection and application in different samples, which demonstrates a rapid auxiliary tool for high-affinity antibody secreted-hybridoma screening and an excellent alternative method for high-quality antibody application.

Development of a Trivalent Construct Omp18/AhpC/FlgH Multi Epitope Peptide Vaccine Against Campylobacter jejuni.

Campylobacter jejuni (C. jejuni) is one of the major pathogens contributing to the enteritis in humans. Infection can lead to numerous complications, including but not limited to Guillain-Barre syndrome, reactive arthritis, and Reiter's syndrome. Over the past two decades, joint efforts have been made toward developing a proper strategy of limiting the transmission of C. jejuni to humans. Nevertheless, except for biosecurity measures, no available vaccine has been developed so far. Judging from the research findings, Omp18, AhpC outer membrane protein, and FlgH flagellin subunits of C. jejuni could be adopted as surface protein antigens of C. jejuni for screening dominant epitope thanks to their strong antigenicity, expression of varying strains, and conservative sequence. In this study, bioinformatics technology was adopted to analyze the T-B antigenic epitopes of Omp18, AhpC, and FlgH in C. jejuni strain NCTC11168. Both ELISA and Western Blot methods were adopted to screen the dominant T-B combined epitope. GGS (GGCGGTAGC) sequence was adopted to connect the dominant T-B combined epitope peptides and to construct the prokaryotic expression system of tandem repeats of antigenic epitope peptides. The mouse infection model was adopted to assess the immunoprotective effect imposed by the trivalent T-B combined with antigen epitope peptide based on Omp18/AhpC/FlgH. In this study, a tandem epitope AhpC-2/Omp18-1/FlgH-1 was developed, which was composed of three epitopes and could effectively enhance the stability and antigenicity of the epitope while preserving its structure. The immunization of BALB/c mice with a tandem epitope could induce protective immunity accompanied by the generation of IgG2a antibody response through the in vitro synthesis of IFN-γ cytokines. Judging from the results of immune protection experiments, the colonization of C. jejuni declined to a significant extent, and it was expected that AhpC-2/Omp18-1/FlgH-1 could be adopted as a candidate antigen for genetic engineering vaccine of C. jejuni MAP.