Additional TCRV beta primers and minor method modifications improve detection of clonal T-cell populations by RT-PCR

• Published in: Molecular pathology Vol. 50; no. 1; pp. 53 - 55
• Main Authors: Lynas, C, Howe, D
• Format: Journal Article
• Language: English
• Published: London BMJ Publishing Group Ltd and Association of Clinical Pathologists 01-02-1997; BMJ
• Subjects: Biological and medical sciences; Clone Cells - immunology; DNA, Complementary - genetics; Electrophoresis, Polyacrylamide Gel; Hot Temperature; Immunoglobulin Variable Region - genetics; Investigative techniques, diagnostic techniques (general aspects); Medical sciences; Miscellaneous. Technology; Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques; Polymerase Chain Reaction - methods; Receptors, Antigen, T-Cell, alpha-beta - genetics; T-Lymphocyte Subsets - immunology; Polymerase chain reaction; Biochemical analysis; Cell line; Beta-Peptide chain; T-Lymphocyte; Molecular biology; Clone; Biological receptor
• ISSN: 1366-8714; 1472-4154
• DOI: 10.1136/mp.50.1.53

The TCRV beta RT-PCR method for detection of clonal populations of T cells which we described previously could not detect clones that used certain variable (V) beta region families. V beta 2, 4, 8.3, and 18 had insufficient homology with the original consensus V region primer. Two new primers have been designed which work well and are able to amplify from V beta families previously undetectable by this RT-PCR. In addition, minor alterations to the cDNA synthesis and gel analysis of the PCR products make the results even easier to interpret. All the Diversity/Joining (D/J) region primer combinations except for D2/J2 have been omitted, and terminating the reverse transcription by heating prior to PCR greatly improves amplification with these primers. Use of 8% and/or 10% polyacrylamide gels increases clarity. Inclusion of the modifications described will reduce false reporting of patients as having a polyclonal T-cell population.