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

ABSTRACT: 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.

SUBMITTER: Lynas C

PROVIDER: S-EPMC379580 | biostudies-other | 1997 Feb

REPOSITORIES: biostudies-other

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

Lynas C C Howe D D

Molecular pathology : MP 19970201 1

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 produ ...[more]

PMID: 9208815

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