title: Relative expression of γδ T-cell receptor gene families detected by RT-PCR and capillary electrophoresis.
abstract: γδ T cells are intensively studied because their function in infection, allergy, autoimmune disease, cancer and post-transplant period is not yet fully understood. PCR-based techniques were established to study the γ variable (Vγ) and δ variable (Vδ) gene families. PCR product evaluation is routinely carried out by Southern blot analysis or the third complementarity-determining region spectratyping, but a fast and simple assessment of Vγ and Vδ gene family expression is missing. The aim of our study was to test capillary electrophoresis as a potential method for evaluating the composition of the γδ T-cell population. This report provides optimized PCR conditions for γδ T-cell receptor amplification. Further, it describes the utilization of capillary electrophoresis in the Agilent 2100 Bioanalyzer to evaluate the relative expression of Vγ and Vδ gene families after their amplification. An application of the methodology to peripheral blood mononuclear cell samples from patients during haemato-oncological treatment is shown. The described methodology is fast and simple to operate and is therefore suitable as a first screening of the γδ T-cell population composition in tissues of interest.
authors.affiliation: Centre for Biomedical Image Analysis, Faculty of Informatics, Masaryk University, Botanická 68a, 60200 Brno, Czech Republic.
authors.forename: L
authors.initials: L
authors.lastname: Tesarova
authors.forename: I
authors.initials: I
authors.lastname: Koutna
authors.forename: M
authors.initials: M
authors.lastname: Dvorakova
authors.forename: M
authors.initials: M
authors.lastname: Klabusay
mesh_terms: D000328:Adult; D016022:Case-Control Studies; D019075:Electrophoresis, Capillary / Q000379:methods*; D005260:Female; D019337:Hematologic Neoplasms / Q000235:genetics / Q000473:pathology; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D005810:Multigene Family; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D016692:Receptors, Antigen, T-Cell, gamma-delta / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism; D013997:Time Factors
chemical_list: D016692:Receptors, Antigen, T-Cell, gamma-delta