title: Evaluation of EBV transformation of human memory B-cells isolated by FACS and MACS techniques.
abstract: Several studies have been performed to develop effective neutralizing monoclonal antibodies. The Epstein-Barr virus (EBV) can efficiently immortalize B-cells to establish lymphoblastoid cell lines (LCL) and so it has been used extensively for transformation of B-cells to produce and secrete immunoglobulin. The present study addressed the effect of TLR7/8 agonist (R848), feeder cells layer and ﬂuorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS) cell separation methods on the transformation efficiency of antibody-producing memory B-cells. For these studies, the antigen used for analyses of antibody formation was the tetanus neurotoxin (TeNT) derived from Clostridium tetani. The results here showed that employing an HFFF.PI6 feeder cell layer, R848 agonist and FACS-mediated purification of memory B-cells led to increased transformation efficiency. Altogether, the effects of the R848 and the feeder cells provided an efficient method for EBV transformation of human B-cells. Moreover, there was an advantage in using FACS sorting of B-cells over the MACS method in the context of EBV transformation and immortalization of precursors of antigen-specific B-cells.
authors.affiliation: a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;
authors.forename: Sanam
authors.initials: S
authors.lastname: Sadreddini
authors.affiliation: a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;|b Department of Immunology, School of Public Health , Tehran University of Medical Sciences , Tehran , Iran ;
authors.forename: Farhad
authors.initials: F
authors.lastname: Jadidi-Niaragh
authors.affiliation: c Monoclonal Antibody Research Center, Avicenna Research Institute , ACECR , Tehran , Iran ;
authors.forename: Vahid
authors.initials: V
authors.lastname: Younesi
authors.affiliation: a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;|d Department of Immunology, School of Medicine , Tabriz University of Medical Sciences , Tabriz , Iran.
authors.forename: Tala
authors.initials: T
authors.lastname: Pourlak
authors.affiliation: a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;
authors.forename: Amir
authors.initials: A
authors.lastname: Afkham
authors.affiliation: b Department of Immunology, School of Public Health , Tehran University of Medical Sciences , Tehran , Iran ;|c Monoclonal Antibody Research Center, Avicenna Research Institute , ACECR , Tehran , Iran ;
authors.forename: Fazel
authors.initials: F
authors.lastname: Shokri
authors.affiliation: a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;|d Department of Immunology, School of Medicine , Tabriz University of Medical Sciences , Tabriz , Iran.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Yousefi
mesh_terms: D000917:Antibody Formation; D001402:B-Lymphocytes / Q000276:immunology* / Q000821:virology; D002461:Cell Line, Transformed; D002471:Cell Transformation, Neoplastic; D003017:Clostridium tetani / Q000276:immunology*; D005434:Flow Cytometry / Q000379:methods*; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007156:Immunologic Memory; D018189:Immunomagnetic Separation / Q000379:methods*; D008666:Metalloendopeptidases / Q000276:immunology; D054448:Precursor Cells, B-Lymphoid / Q000276:immunology* / Q000821:virology; D013744:Tetanus Toxin / Q000276:immunology; D051199:Toll-Like Receptor 7 / Q000037:antagonists & inhibitors; D051200:Toll-Like Receptor 8 / Q000037:antagonists & inhibitors
chemical_list: D007093:Imidazoles; C495348:TLR7 protein, human; C495349:TLR8 protein, human; D013744:Tetanus Toxin; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; C035979:tetanospasmin; D008666:Metalloendopeptidases; C402365:resiquimod
keywords: B-cell transformation; Epstein–Barr virus; FACS; MACS; monoclonal antibody; tetanus