An improved methodology to detect human T cell receptor beta variable family gene expression patterns.
Comprehensive gene expression analysis of the T cell receptor repertoire of an individual can be very useful in evaluating the immune response in a variety of conditions. Antibody-based analysis methods can detect approximately 60% of the human T cell receptor beta variable (TCRBV) proteins, while gene expression analysis, primarily through employment of the polymerase chain reaction (PCR), has had somewhat greater success in the detection of additional TCRBV families. Many of these previous PCR methods, however, have been unable to detect all 91 alleles of the human TCRBV genes. This is primarily due to either deficiencies in the amplification of all of the variable beta families, subfamilies, and alleles, or the prior lack of a systematic classification of the TCR variable family gene segment sequences. We describe here a real-time reverse transcription polymerase chain reaction-based method, which allows efficient automation and integration of amplification, detection, and analysis with sequence-specific detection of all T cell receptor beta variable gene families, subfamilies, and alleles. This method, which in itself contributes significant improvements over existing technologies through its comprehensiveness and efficiency, also functions independently of variables such as sample source and sample processing and has the ability to run on multiple real-time PCR platforms, affording one the implementation of personal preferences.
Brewer
Jamie Leigh
JL
Department of Microbiology, Immunology, and Cell Biology, West Virginia University, PO Box 9177, Morgantown, WV 26506-9177, USA. jbrewer@hsc.wvu.edu
Ericson
Solveig Gronning
SG
D017931:DNA Primers; D005470:Fluorometry / Q000379:methods; D020869:Gene Expression Profiling / Q000379:methods*; D014644:Genetic Variation; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D006801:Humans; D005810:Multigene Family; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000096:biosynthesis; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000096:biosynthesis* / Q000235:genetics*
D017931:DNA Primers; D012333:RNA, Messenger; D016693:Receptors, Antigen, T-Cell, alpha-beta