Construction of a Recombinant OmpC Dominant Epitope-Based Vaccine Against Escherichia coli and Evaluation of Its Immunogenicity and Protective Immunity

Liu, Xiang; Chen, Chunlin; Chen, Chen; Ding, Rui; Marslin, Gregory

doi:10.5812/jjm.55652

Cited by 3 publications

(7 citation statements)

References 35 publications

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“…At an OD 600 value of 0.5, IPTG was added to a final concentration of 0.5 mmol/L; the strains were then cultured at 37°C for 5 h. The expression of OmpA was assessed by SDS-PAGE electrophoresis. The OmpA protein was loaded onto a Ni-affinity chromatography column and purified using a Ni-NTA flowresin method (Sigma-Aldrich, USA) [12].…”

Section: Prokaryotic Expression and Purification Of Ompa Proteinmentioning

confidence: 99%

“…OmpA protein and the control of PBS solution were emulsified with Freund's Complete Adjuvant (Sigma-Aldrich, USA) for the first immunization. After 14 days, the mice were boosted using Freund's incomplete adjuvant (Sigma-Aldrich, USA) [12]. After 7 days, a third immunization was performed.…”

Section: Preparation Of Mouse Anti-ompa Polyclonal Antiserummentioning

confidence: 99%

“…Then, the membrane was incubated with horseradish peroxidase-conjugated anti-mouse secondary antibody (TaKaRa, Japan). The NC membrane was then incubated with a DAB solution (Sigma-Aldrich, USA) coloration system to visualize bands [12]. OmpA serum specificity was determined according to the color of NC membrane bands.…”

Section: Specificity and Titer Detection Of Ompa Protein Antiserummentioning

confidence: 99%

“…Coloration liquid was added to every tube to avoid light reaction. After a stop solution was added to each tube, a microplate reader was used to detect the absorbance value at OD 450 [12].…”

Section: Interaction Between Ompa Antiserum and Pathogens Of Animal Mmentioning

confidence: 99%

“…The immune protection rates were expressed as a formula of 1 -(OmpA immunity mortality/non-OmpA immunity mortality) × 100%. The statistical software package Social Science (SPSS) was used for statistical significance analysis [11,12].…”

Section: Immune Protective Function Of Ompa Proteinmentioning

confidence: 99%

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Immunoprotective evaluation of <I>Escherichia coli</I> outer membrane protein A against the main pathogens of animal mastitis

Chen¹,

Wu²,

Na³

et al. 2020

Trop. J. Pharm Res

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Purpose: To evaluate prokaryotic expression of the Escherichia coli (E. coli) outer membrane protein A (OmpA) and its immunoprotective function against the main pathogens of animal mastitis.Methods: A molecular cloning method was used to develop a prokaryotic strain expressing OmpA protein, which was purified by Ni-affinity chromatography. Polyclonal antiserum was generated in mice immunized with OmpA protein. Enzyme-linked immunosorbent assay (ELISA) and western blotting were used to determine the titer and verify anti-OmpA serum specificity, respectively. Interaction between OmpA antiserum and main pathogens of animal mastitis was verified by ELISA and a pull-down method. The immune protective function of OmpA protein was evaluated in mice challenged with pathogens of animal mastitis. Optimal fermentation conditions to produce OmpA protein were determined by the L9(34) orthogonal test.Results: A prokaryotic strain expressing OmpA protein was developed, and purified OmpA was used to develop a mouse polyclonal antibody. The anti-OmpA serum exhibited high specificity and a titer of 1:1600. Anti-OmpA serum directly interacted with E. coli and Staphylococcus aureus (S. aureus). OmpA demonstrated a significant immune protective function of 58.33 % against E. coli and 46.15 % against S. aureus. The optimal conditions for expressing fermentation OmpA were a strain absorbance of 0.5 at a wavelength of 600 nm, IPTG final concentration of 0.3 mmol/L, induction time of 12 h, and induction temperature of 28 °C.Conclusion: OmpA possesses selective immunogenicity and a significant immune protective effect against the main pathogens of animal mastitis. The results suggest that OmpA may potentially be used as a vaccine for animal mastitis. Keywords: E. coli, OmpA protein, Immunoprotection, Animal mastitis, Protein fermentation

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Section: Prokaryotic Expression and Purification Of Ompa Proteinmentioning

confidence: 99%

Section: Preparation Of Mouse Anti-ompa Polyclonal Antiserummentioning

confidence: 99%

Section: Specificity and Titer Detection Of Ompa Protein Antiserummentioning

confidence: 99%

Section: Interaction Between Ompa Antiserum and Pathogens Of Animal Mmentioning

confidence: 99%

Section: Immune Protective Function Of Ompa Proteinmentioning

confidence: 99%

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Immunoprotective evaluation of <I>Escherichia coli</I> outer membrane protein A against the main pathogens of animal mastitis

Chen¹,

Wu²,

Na³

et al. 2020

Trop. J. Pharm Res

Self Cite

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Epitope Prediction by Novel Immunoinformatics Approach: A State-of-the-art Review

Raoufi

Hemmati

Eftekhari

et al. 2019

Int J Pept Res Ther

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Immunoinformatics is a science that helps to create significant immunological information using bioinformatics softwares and applications. One of the most important applications of immunoinformatics is the prediction of a variety of specific epitopes for B cell recognition and T cell through MHC class I and II molecules. This method reduces costs and time compared to laboratory tests. In this state-of-the-art review, we review about 50 papers to find the latest and most used immunoinformatic tools as well as their applications for predicting the viral, bacterial and tumoral structural and linear epitopes of B and T cells. In the clinic, the main application of prediction of epitopes is for designing peptide-based vaccines. Peptide-based vaccines are a considerably potential alternative to low-cost vaccines that may reduce the risks related to the production of common vaccines.

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Evaluation of Immunogenicity, Protective Immunity on Aquaculture Pathogenic Vibrio and Fermentation of Vibrio alginolyticus Flagellin FlaC Protein

Chen

Kang

et al. 2019

IRAN J BIOTCH

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Background: Vibrio are the main pathogenic bacteria in aquaculture. The flagellin protein C (FlaC) of Vibrio alginolyticus has good immunogenicity and the prospect of potential application in a vaccine. Objectives: We aimed to evaluate the immunogenicity, protective immunity, and prokaryotic expression fermentation of V. alginolyticus FlaC protein for the vaccine in aquaculture. Material and Methods: A molecular cloning method was used to construct the expression strain of FlaC protein, and the protein was purified with Ni-affinity chromatography. Polyclonal antiserum was prepared via mice immunized with the FlaC protein. The Western blot and enzyme-linked immunosorbent assay (ELISA) were used to check the specificity and titre of the antiserum. ELISA and pull-down assay detected the interaction between FlaC protein antiserum and Vibrio. The immune protection function of FlaC protein was detected with mice actively immunized with FlaC protein and challenged by V. alginolyticus and V. parahaemolyticus. The optimal expression conditions for FlaC protein were detected using an L9(3 4 ) orthogonal design model. Results:The expression strain of FlaC protein was obtained successfully, and purified FlaC protein was prepared using a mice polyclonal antibody. The FlaC protein antiserum held a high specificity, and the titre was 1:3200. The antiserum directly interacted with V. alginolyticus and V. parahaemolyticus, and the FlaC protein demonstrated a significant immune protection function (50%) against V. alginolyticus infection and some immune protection function (41.66%) against V. parahaemolyticus. The optimal expression conditions for FlaC protein included a strain OD600 value of 0.8, final isopropyl-β-d-thiogalactoside (IPTG) concentration of 0.1 mmol/L, an inducing time of 8 hours, and an inducing temperature of 28°C. Conclusions: This study showed that the FlaC protein possesses a significant immunogenicity and immune protection effect and obtained the optimal fermentation conditions. It is expected to be a potential vaccine against V. alginolyticus and V. parahaemolyticus.

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Construction of a Recombinant OmpC Dominant Epitope-Based Vaccine Against Escherichia coli and Evaluation of Its Immunogenicity and Protective Immunity

Cited by 3 publications

References 35 publications

Immunoprotective evaluation of <I>Escherichia coli</I> outer membrane protein A against the main pathogens of animal mastitis

Immunoprotective evaluation of <I>Escherichia coli</I> outer membrane protein A against the main pathogens of animal mastitis

Epitope Prediction by Novel Immunoinformatics Approach: A State-of-the-art Review

Evaluation of Immunogenicity, Protective Immunity on Aquaculture Pathogenic Vibrio and Fermentation of Vibrio alginolyticus Flagellin FlaC Protein

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