A novel method for making human monoclonal antibodies.
We have developed a B cell immortalization method for low B cell numbers per well using simultaneous B cell stimulation by CpG2006 and B cell infection by Epstein-Barr virus (EBV), followed by an additional CpG2006 and interleukin-2 (IL-2) stimulus. Using this method, immunoglobulin G (IgG)-producing immortalized B cell lines were generated from peripheral blood IgG(+)CD22(+) B cells with an efficiency of up to 83%. Antibody can already be obtained from the culture supernatant after 3-4 weeks. Moreover, clonality analysis demonstrated monoclonality in 87% of the resulting immortalized B cell lines. Given the high immortalization efficiency and monoclonality rate, evidence is provided that no further subcloning is necessary. An important application of this B cell immortalization method is the characterization of (autoreactive) antibodies from patients with autoimmune disease. This could eventually lead to the identification of new autoantigens, disease markers or targets for therapy.
Fraussen
J
J
Hasselt University, Biomedical Research Institute and Transnationale Universiteit Limburg, School of Life Sciences, Diepenbeek, Belgium.
Vrolix
K
K
Martinez-Martinez
P
P
Losen
M
M
Meulemans
E
E
De Baets
M H
MH
Stinissen
P
P
Somers
V
V
D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D001402:B-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism* / Q000473:pathology / Q000821:virology; D018929:Cell Culture Techniques / Q000379:methods; D002472:Cell Transformation, Viral / Q000187:drug effects / Q000276:immunology; D002999:Clone Cells; D004247:DNA / Q000032:analysis; D020031:Epstein-Barr Virus Infections / Q000276:immunology*; D004854:Herpesvirus 4, Human / Q000276:immunology* / Q000472:pathogenicity; D006801:Humans; D006825:Hybridomas; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D007376:Interleukin-2 / Q000494:pharmacology*; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology*; D051918:Sialic Acid Binding Ig-like Lectin 2 / Q000096:biosynthesis
D000911:Antibodies, Monoclonal; C513533:CpG ODN 2006; D007074:Immunoglobulin G; D007376:Interleukin-2; D009838:Oligodeoxyribonucleotides; D051918:Sialic Acid Binding Ig-like Lectin 2; D004247:DNA