Title:In Vitro Evaluation of Immunogenicity of Recombinant OMP25 Protein Obtained from Endemic Brucella abortus Biovar 3 as Vaccine Candidate Molecule Against Animal Brucellosis
Volume: 28
Issue: 10
Author(s): Tuğba Atabey, Tayfun Acar, Serap Derman, Emel Ordu, Ayşegül Erdemir, Pakize Neslihan Taşlı, Günseli Kurt Gür, Fikrettin Şahin, Medine Güllüce and Tülin Arasoğlu*
Affiliation:
- Molecular Biology and Genetics Department, Faculty of Arts and Sciences, Yildiz Technical University, Istanbul,Turkey
Keywords:
Brucellosis, Brucella abortus, vaccine, OMP25, recombinant protein, fusion protein.
Abstract:
Background: Brucellosis is a zoonotic disease that causes serious economic losses due
to factors, such as miscarriages and decreased milk yield in animals. Existing live vaccines have
some disadvantages, so effective vaccines need to be developed with new technological approaches.
Objective: The primary objectives of this study were the expression and purification of recombinant
Omp25 fusion protein from B. abortus, and the evaluation of the effect of the Omp25 protein
on cell viability and inflammatory response.
Methods: The omp25 gene region was amplified by a polymerase chain reaction and cloned into a
Pet102/D-TOPO expression vector. The protein expression was carried out using the prokaryotic
expression system. The recombinant Omp25 protein was purified with affinity chromatography followed
by GPC (Gel Permeation Chromatography). The MTS assay and cytokine-release measurements
were carried out to evaluate cell viability and inflammatory response, respectively.
Results: It was determined that doses of the recombinant Omp25 protein greater than 0.1 μg/mL
are toxic to RAW cells. Doses of 1 μg/mL and lower significantly increased inflammation due to
Nitric Oxide (NO) levels. ELISA results showed that IFN-γ was produced in stimulated RAW
264.7 cells at a dose that did not affect the viability (0.05 μg/mL). However, IL-12, which is
known to have a dual role in the activation of macrophages, did not show a statistically significant
difference at the same dose.
Conclusion: Studies on cell viability and Th1-related cytokine release suggest Omp25 protein to
be a promising candidate molecule for vaccine development.
