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کاربرد نوع شرط:
- جایگاه : پژوهشی
- مجله: Iranian Journal of Basic Medical Sciences
- نوع مقاله: Journal Article
- کلمات کلیدی: ELISA,vaccine,Brucella melitensis,Fusion protein,Immune reactivity
- چکیده:
- چکیده انگلیسی: Objective(s): Brucella spp. are facultative intracellular pathogens which can stay alive and multiply in professional and nonprofessional phagocytes. Immunity against Brucella melitensis involves antigen-specific CD4+ and CD8+ T-cells activation and humoral immune responses. Due to negative aspects of live attenuated vaccines, much attention has been focused on finding Brucella-protective antigens to introduce them as potential subunit vaccine candidates.
Materials and Methods: A chimeric gene encoding trigger factor (TF), Omp3148-74 and BP2687-111 fragments (TOB) from B. melitensis was successfully cloned, expressed in Escherichia coliBL21-DE3 and purified by Ni-NTA agarose column. Antibodies to recombinant TOB (rTOB) have been investigated in Brucella-infected human sera and a pool serum prepared from B. melitensis-vaccinated rabbits.
Results: Our results showed that the immunized rabbit pool serum strongly reacted with rTOB. In addition, antibodies against rTOB were detectable in 76.5% of sera obtained from infected patients.
Conclusion: These findings suggest that rTOB may provide a potential immunogenic candidate which could be considered in future vaccine studies.- انتشار مقاله: 13-02-1394
- نویسندگان: Jafar Amani,Amir Ghasemi,Reza Ranjbar,Mahdi Shabani,Mahdi Zandemami,Reza Golmohammadi
- مشاهده
- جایگاه : پژوهشی
- مجله: Iranian Journal of Biotechnology
- نوع مقاله: Journal Article
- کلمات کلیدی: Purification,Protein expression,Polyclonal antibody,FCRL,His-tag
- چکیده:
- چکیده انگلیسی: Background: The Fc receptor like (FCRL) molecules belong to the immunoglobulin (Ig) superfamily with potentially immunoregulatory function. Among the FCRL family FCRL2 and 4 are predominantly expressed on memory B cells and FCRL1 is a pan- B cell marker. To date, no ligand has been identifid for the human FCRL1, 2 and 4 molecules. Objectives: Cloning, expression, purifiation and structural analysis of the extracellular domain of human FCRL1, 2 and 4 proteins. Materials and Methods: In this study, the extracellular part of human FCRL1, 2 and 4 were subcloned into prokaryotic expression vectors pET-28b (+) and transformed into BL21-DE3 E.coli strain. Protein expression was optimized by fie adjustments such as induction time, incubation temperature and expression hosts. Recombinant FCRL proteins were purifid by metal affity chromatography using Ni-NTA resin. Purifid FCRL proteins were further characterized by SDS-PAGE and immunoblotting using His-tag and FCRL specifi polyclonal antibodies. Results: Our results demonstrated that FCRL1, 2 and 4 were successfully expressed in pET-28b (+) vector. Optimization of the expression procedure showed that IPTG induction at OD600 = 0.9 and overnight incubation at 37˚C resulted in the highest expression levels of FCRL proteins ranging from approximately 15% (FCRL1) to 25% (FCRL2 and 4) of the total bacterial lysate proteins. Conclusions: These purifid recombinant proteins are potentially a valuable tool for investigating the immunoregulatory function of FCRL molecules and the production of specifi mAbs for immunotherapeutic interventions.
- انتشار مقاله: 04-10-1391
- نویسندگان: Mahdi Shabani,Azam Hemmati,Mahdi Zandemami,Jalal Khoshnoodi,Mahmood Jeddi-Tehrani,Hodjatallah Rabbani,Zahra Amirghofran,Fazel Shokri
- مشاهده