Author/Authors :
Delbaz، Seyed Ali نويسنده Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, IR Iran , , Siadat، Seyed Davar نويسنده , , Aghasadeghi، Mohammad Reza نويسنده , , Piryaie، Mohamad نويسنده Department of Genetics, Faculty of Medical Sciences, Shahid Beheshti University of Medical Sciences, Tehran, IR Iran , , Najar Peerayeh، Shahin نويسنده , , Mousavi، Seyed Fazlollah نويسنده Department of Microbiology, Pasteur Institute of Iran, Tehran, IR Iran , , Moshiri، Arfa نويسنده , , Sadat، Seyed Mehdi نويسنده , , Zangeneh، Mehrangiz نويسنده Department of Infectious Diseases, Tehran Medical Branch, Islamic Azad University, Tehran, IR Iran , , Nejati، Mehdi نويسنده Department of Microbiology, Pasteur Institute of Iran, Tehran, IR Iran , , Kashanizadeh، Nafiseh نويسنده Department of Gynocology, Baghiat-Alah University, Tehran, IR Iran , , Bouzari، Saied نويسنده Department of Molecular Biology, Pasteur Institute of Iran, Tehran, IR Iran ,
Abstract :
Background: Neisseria meningitidis Serogroup A, is a major cause of bacterial meningitidis outbreaks in Africa and the Middle East. While polysaccharide vaccines have been available for many years, these vaccines have many disadvantages including the induction of T-cell independent responses which do not induce memory responses.
Objectives: Thus to overcome this problem, in this research outer membrane vesicle (OMV) containing PorA was extracted and evaluated by biological and immunological methods.
Materials and Methods: OMVs were extracted with deoxycholate and EDTA, and purification was performed by sequential ultracentrifugation. Physicochemical properties of extracted OMVs were analyzed by electron microscopy and SDS-PAGE. The toxicity of LPS content in its was assayed by LAL test. The Presence of PorA as a major component of OMV was confirmed by western blot. To study antibodies synthesis after immunization with OMV, ELISA method was used. Also serum bactericidal assay (SBA) was performed to determine the serum bactericidal activity against N.meningitidis serogroup A.
Results: The results revealed that the content of protein extracted was 0.1mg/ml. The electron microscopy showed that intactness of the vesicle in these preparation ranged more than 70%. The SDS-PAGE showed that PorA as a major immunological part of outer membrane vesicle was located in 35-40kDa. LAL test showed that the endotoxin activity was around 126EU/ml which is safe for using. The ELISA test revealed that the IgG total titer was elevated after the first injection. SBA indicates that bactericidal antibodies rise after the second dose of booster.
Conclusions: The results showed that the extracted OMVs were conformationally stable, and there were no pyrogenic determinants in OMV. Also the results showed that the OMV elicited high level of specific antibodies against N. meningitidis serogroup A. These results indicate that the OMV obtained here, can be used as a meningococcal vaccine after further investigation.
