Author/Authors :
-، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran
and
Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, IR Iran Shabani, Mahdi , -، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran Hemmati, Azam , -، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran Zandemami, Mahdi , -، - نويسنده Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, IR Iran Khoshnoodi, Jalal , -، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran Jeddi-Tehrani, Mahmood , -، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran Rabbani, Hodjatallah , -، - نويسنده Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, IR Iran Amirghofran, Zahra , -، - نويسنده Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, IR Iran Shokri, Fazel
Abstract :
Background: The Fc receptor like (FCRL) molecules belong to the immunoglobulin (Ig) superfamily with potentially immunoregulatoryfunction. Among the FCRL family FCRL2 and 4 are predominantly expressed on memory B cells and FCRL1 is a pan- B cell marker. To date, noligand 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 vectorspET-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-NTAresin. Purifid FCRL proteins were further characterized by SDS-PAGE and immunoblotting using His-tag and FCRL specifi polyclonalantibodies.Results: Our results demonstrated that FCRL1, 2 and 4 were successfully expressed in pET-28b (+) vector. Optimization of the expressionprocedure showed that IPTG induction at OD600 = 0.9 and overnight incubation at 37˚C resulted in the highest expression levels of FCRLproteins 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 ofFCRL molecules and the production of specifi mAbs for immunotherapeutic interventions.
