Limits...
Immunogenicity of a fusion protein comprising coli surface antigen 3 and labile B subunit of enterotoxigenic Escherichia coli.

Alerasol M, Mousavi Gargari SL, Nazarian S, Bagheri S - Iran. Biomed. J. (2014)

Bottom Line: Pre-treatment of the ETEC cells with anti-sera significantly decreased their adhesion to Caco-2 cells.The results indicated the efficacy of the recombinant chimeric protein as an effective immunogen inducing strong humoral response.The designated chimer would be an interesting prototype for a vaccine and worthy of further investigation.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Biology, Shahed University, Tehran, Iran. slmousavi@shahed.ac.ir.

ABSTRACT

Background: Enterotoxigenic Escherichia coli (ETEC) strains are the major causes of diarrheal disease in humans and animals. Colonization factors and enterotoxins are the major virulence factors in ETEC pathogenesis. For the broad-spectrum protection against ETEC, one could focus on colonization factors and non-toxic heat labile as a vaccine candidate.

Methods: A fusion protein is composed of a major fimbrial subunit of coli surface antigen 3, and the heat-labile B subunit (LTB) was constructed as a chimeric immunogen. For optimum level expression of protein, the gene was synthesized with codon bias of E. coli. Also, recombinant protein was expressed in E. coli BL21DE3. ELISA and Western tests were carried out for determination of antigen and specificity of antibody raised against recombinant protein in animals. The anti-toxicity and anti-adherence properties of the immune sera against ETEC were also evaluated.

Results: Immunological analyses showed the production of high titer of specific antibody in immunized mice. The built-in LTB retains native toxin properties which were approved by GM1 binding assay. Pre-treatment of the ETEC cells with anti-sera significantly decreased their adhesion to Caco-2 cells.

Conclusion: The results indicated the efficacy of the recombinant chimeric protein as an effective immunogen inducing strong humoral response. The designated chimer would be an interesting prototype for a vaccine and worthy of further investigation.

Show MeSH

Related in: MedlinePlus

Determination of inhibition of the binding of heat labile to GM1 using GM1 ELISA assay. Each graph shows the mean OD ± SD in three independent experiments. GM1(1), GM1 binding to the native toxin as positive control; GM1(2), neutralized toxin by CstH:LTB anti-serum lost GM1 binding ability, and control, no GM1 coated in control strips
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4225060&req=5

Figure 6: Determination of inhibition of the binding of heat labile to GM1 using GM1 ELISA assay. Each graph shows the mean OD ± SD in three independent experiments. GM1(1), GM1 binding to the native toxin as positive control; GM1(2), neutralized toxin by CstH:LTB anti-serum lost GM1 binding ability, and control, no GM1 coated in control strips

Mentions: Activity inhibition assay of toxin. Biological activity and toxin neutralizing efficacy of the CstH:LTB-specific antibodies were tested by GM1-binding inhibition assay. Incubation of toxin with CstH:LTB antibody resulted in toxin neutralization and its prevention to bind to GM1 (Fig. 6). The sera from control mice or PBS did not show such effect. Induction of fluid accumulation by toxin was studied in rabbit ileal loops. The volume of secretion for each ileal loop was measured after excision. The fluid accumulation was not observed in rabbit ileal loops 18 h post infection with ETEC treated with the immunized serum. The volume per length ratios of ileal loops (ml/cm) of the rabbit injected with ETEC + anti-chimeric protein, ETEC, and PBS were recorded 0.81, 2.23, and 0.51, respectively.


Immunogenicity of a fusion protein comprising coli surface antigen 3 and labile B subunit of enterotoxigenic Escherichia coli.

Alerasol M, Mousavi Gargari SL, Nazarian S, Bagheri S - Iran. Biomed. J. (2014)

Determination of inhibition of the binding of heat labile to GM1 using GM1 ELISA assay. Each graph shows the mean OD ± SD in three independent experiments. GM1(1), GM1 binding to the native toxin as positive control; GM1(2), neutralized toxin by CstH:LTB anti-serum lost GM1 binding ability, and control, no GM1 coated in control strips
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225060&req=5

Figure 6: Determination of inhibition of the binding of heat labile to GM1 using GM1 ELISA assay. Each graph shows the mean OD ± SD in three independent experiments. GM1(1), GM1 binding to the native toxin as positive control; GM1(2), neutralized toxin by CstH:LTB anti-serum lost GM1 binding ability, and control, no GM1 coated in control strips
Mentions: Activity inhibition assay of toxin. Biological activity and toxin neutralizing efficacy of the CstH:LTB-specific antibodies were tested by GM1-binding inhibition assay. Incubation of toxin with CstH:LTB antibody resulted in toxin neutralization and its prevention to bind to GM1 (Fig. 6). The sera from control mice or PBS did not show such effect. Induction of fluid accumulation by toxin was studied in rabbit ileal loops. The volume of secretion for each ileal loop was measured after excision. The fluid accumulation was not observed in rabbit ileal loops 18 h post infection with ETEC treated with the immunized serum. The volume per length ratios of ileal loops (ml/cm) of the rabbit injected with ETEC + anti-chimeric protein, ETEC, and PBS were recorded 0.81, 2.23, and 0.51, respectively.

Bottom Line: Pre-treatment of the ETEC cells with anti-sera significantly decreased their adhesion to Caco-2 cells.The results indicated the efficacy of the recombinant chimeric protein as an effective immunogen inducing strong humoral response.The designated chimer would be an interesting prototype for a vaccine and worthy of further investigation.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Biology, Shahed University, Tehran, Iran. slmousavi@shahed.ac.ir.

ABSTRACT

Background: Enterotoxigenic Escherichia coli (ETEC) strains are the major causes of diarrheal disease in humans and animals. Colonization factors and enterotoxins are the major virulence factors in ETEC pathogenesis. For the broad-spectrum protection against ETEC, one could focus on colonization factors and non-toxic heat labile as a vaccine candidate.

Methods: A fusion protein is composed of a major fimbrial subunit of coli surface antigen 3, and the heat-labile B subunit (LTB) was constructed as a chimeric immunogen. For optimum level expression of protein, the gene was synthesized with codon bias of E. coli. Also, recombinant protein was expressed in E. coli BL21DE3. ELISA and Western tests were carried out for determination of antigen and specificity of antibody raised against recombinant protein in animals. The anti-toxicity and anti-adherence properties of the immune sera against ETEC were also evaluated.

Results: Immunological analyses showed the production of high titer of specific antibody in immunized mice. The built-in LTB retains native toxin properties which were approved by GM1 binding assay. Pre-treatment of the ETEC cells with anti-sera significantly decreased their adhesion to Caco-2 cells.

Conclusion: The results indicated the efficacy of the recombinant chimeric protein as an effective immunogen inducing strong humoral response. The designated chimer would be an interesting prototype for a vaccine and worthy of further investigation.

Show MeSH
Related in: MedlinePlus