Improved Polymerase Chain Reaction Detection of Clonally Rearranged T-Cell Receptor β Chain Genes

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Abstract
A new method for the detection of all known possible rearrangements at the variable (V), diversity (D), and joining (J) segments of the T-cell receptor p chain (TcR[beta]) gene in tissue DNA extracts is described that involves two polymerase chain reactions (PCRs). The first PCR round (screening PCR) allowed the identification of the J[beta] segment involved in a clonal rearrangement. A J[beta]-primer was used for the second PCR (J[beta]-specific PCR), recognizing the J[beta] segment identified in the screening PCR in combination with a consensus V[beta] primer. This PCR generated prominent and short amplificates suitable for direct sequence analysis because of their low background. Using this approach, clonal TcR[beta] gene rearrangements were able to be demonstrated in all T-cell lines (n = 7) and in all peripheral T-cell lymphomas (n = 33) analyzed. No clonal TcR[beta] gene rearrangements were found in any of the normal tissues studied nor in any B-cell non-Hodgkin lymphomas. This method is applicable to DNA from fresh frozen tissues, and, after the TcR[beta] rearrangement of a patient's malignant T-cell clone has been identified by the screening PCR, DNA can also be detected in follow-up formalin-fixed paraffin-embedded samples by the J[beta]-specific PCR with high sensitivity and specificity. (C) Lippincott-Raven Publishers.

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