Rapid generation of specific antibodies by enhanced homologous recombination.
In the chicken immune system, gene conversion, a type of homologous recombination, primarily contributes to diversification of the immunoglobulin gene. Here, we report on the rapid generation of specific monoclonal antibodies using the chicken DT40 B-cell line undergoing gene conversion. We discovered that the gene conversion frequency at the immunoglobulin locus is increased by treating DT40 cells with a histone deacetylase inhibitor, trichostatin A (TSA), thereby generating diversity at the immunoglobulin locus in the majority of treated cells. This indicates that TSA treatment accelerates the autonomous diversification of surface IgMs on DT40 cells. We took advantage of this effect to select DT40 cells producing specific antibodies with antigen-conjugated magnetic beads. This autonomously diversifying library (ADLib) selection system enables the quick establishment (approximately 1 week from a diversifying library) of various clones producing monoclonal IgMs with enough specificity and affinity for immunological assays, and is applicable to various biotechnologies including rational protein design.
Seo
Hidetaka
H
Genetic Dynamics Research Unit, RIKEN Discovery Research Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Masuoka
Mieko
M
Murofushi
Hiromu
H
Takeda
Shunichi
S
Shibata
Takehiko
T
Ohta
Kunihiro
K
D000818:Animals; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D001402:B-Lymphocytes / Q000276:immunology; D002460:Cell Line; D003509:Cyclohexanecarboxylic Acids / Q000276:immunology; D015322:Gene Rearrangement, B-Lymphocyte; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids / Q000494:pharmacology; D007128:Immunoglobulin Fragments / Q000235:genetics*; D007075:Immunoglobulin M / Q000235:genetics*; D008280:Magnetics; D010047:Ovalbumin / Q000276:immunology; D011995:Recombination, Genetic; D019809:Streptavidin / Q000276:immunology; D014170:Transformation, Genetic
D003509:Cyclohexanecarboxylic Acids; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D007128:Immunoglobulin Fragments; D007075:Immunoglobulin M; C012589:trichostatin A; C045466:IG 10; D010047:Ovalbumin; D019809:Streptavidin