Analysis of cellular phenotype during in vitro immunization of murine splenocytes for generating antigen-specific immunoglobulin

作者：Inagaki Takashi; Yoshimi Tatsunari; Kobayashi Satoshi; Kawahara Masahiro; Nagamune Teruyuki^*

来源：Journal of Bioscience and Bioengineering, 2013, 115(3): 339-345.

DOI：10.1016/j.jbiosc.2012.10.008

• In vitro immunization
• Activation-induced cytidine deaminase
• Germinal center
• Somatic hypermutation
• T cell dependent stimuli

Although various in vitro immunization methods to generate antigen-specific antibodies have been described, a highly effective method that can generate high-affinity immunoglobulins has not yet been reported. Herein, we analyzed a cellular phenotype during in vitro immunization of murine splenocytes for generating antigen-specific immunoglobulins. We identified a combination of T cell-dependent stimuli (IL-4, IL-5, anti-CD38 and anti-CD40 antibodies) plus lipopolysaccharides (LPS) that stimulates antigen-exposed splenocytes in vitro, followed by induction of the cells phenotypically equivalent to germinal center B cells. We also observed that LPS induced high expression levels of mRNA for activation-induced cytidine deaminase. We stimulated antigen-exposed splenocytes, followed by the accumulation of mutations in immunoglobulin genes. From the immunized splenocytes, hybridoma clones secreting antigen-specific immunoglobulins were obtained.

• 出版日期2013-3