IMPROVED POLYMERASE-CHAIN-REACTION DETECTION OF CLONALLY REARRANGED T-CELL RECEPTOR-BETA CHAIN GENES

A new method for the detection of all known possible rearrangements at the variable (V), diversity (D), and joining (J) segments of the T-ce ll receptor beta chain (TcR beta) gene in tissue DNA extracts is descr ibed that involves two polymerase chain reactions (PCRs). The first PC R round (screening PCR) allowed the identification of the J beta segme nt involved in a clonal rearrangement. A J beta-primer was used for th e second PCR (J beta-specific PCR), recognizing the J beta segment ide ntified in the screening PCR in combination with a consensus V beta pr imer. 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 demonstr ated in all T-cell lines (n = 7) and in all peripheral T-cell lymphoma s (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 ly mphomas. This method is applicable to DNA from fresh frozen tissues, a nd, after the TcR beta rearrangement of a patient's malignant T-cell c lone has been identified by the screening PCR, DNA can also be detecte d in follow-up formalin-fixed paraffin-embedded samples by the J beta- specific PCR with high sensitivity and specificity.