Majallah-i Dānishgāh-i ̒Ulūm-i Pizishkī-i Bābul (Sep 2017)
Immunogenicity Evaluation of PLGA Nanoparticles Contains Recombinant CfaB Protein from Enterotoxigenic Escherichia Coli
Abstract
BACKGROUND AND OBJECTIVE: Enterotoxigenic Escherichia coli (ETEC) are the most common causes of diarrhea among children. Colonization factors (CFs) are the major ETEC candidate vaccines. Considering the high antibiotics resistance reports, construction of effective immunogen against bacteria is major goal. Encapsulation of recombinant immunogen in nano carriers such as polymeric PLGA nanparticles, not only protects them from degradation in environmental elements but also provides the effective concentrations of immunogen in targets, hence increasing bioactivity. The aim of this study was to investigate the immunogenic properties of CfaB protein encapsulated in PLGA nanoparticles. METHODS: In this research study, cfaB gene codon optimaztion was performed by OPTIMIZER software. Expression of recombinant protein in E.coli was induced with IPTG. The recombinant protein was purified using affinity chromatography and evaluated by Western Blotting analysis. PLGA nanoparticles containing recombinant protein were prepared by double emulsion method and their structures were characterized by SEM and DLS. 40 BALB/C mice in four group were immunized with nanoparticles and antibody titers were determined by ELISA. The efficiency of antibodies in preventing the attachment of ETEC bacteria to the Caco 2 cells was examined. FINDINGS: The optimized gene had a Codon adaptation index (CAI) of 0.85. Expression of recombinant protein led to the production of CfaB with molecular weight of 18/9 kDa. Yield of protein was 5mg/L. Encapsulation efficiency of recombinant protein in PLGA nanoparticles was 85٪. The average particle size was 170 nm. Immunization of mice induced serum antibody response. Immunized sera decreased %62.8 of ETEC adhesion to Caco2 cells. CONCLUSION: Recombinant CfaB protein encapsulated in PLGA nanoparticles stimulate immunity against ETEC.
