In situ determination of T-cell receptor beta expression patterns

A definitive diagnosis of T-cell lymphoma may be contingent on the rearrangement profile of the T-cell receptor. This is most accurately done by molecular analysis of the beta-chain of the T-cell receptor (TCRβ) by Southern blotting hybridization that requires unfixed tissue. We describe a reverse transcriptase in situ PCR (RT in situ PCR) method that permits the target-specific direct incorporation of the reporter nucleotide into the different transcripts that comprise the TCRβ, using paraffin-embedded, formalin-fixed tissue. Each of the 25 possible Vβ segment rearrangments was documented in three lymph nodes with nonspecific lymphadenitis, with clonal expansion evident in a case of metastatic melanoma. Monoclonal expression was documented in seven tissues diagnostic of a T-cell lymphoma. We analyzed five additional tissues for which a definitive diagnosis of T-cell vs B-cell lymphoma could not be rendered on the basis of histological, immunohistological, and flow cytometric analysis. RT in situ PCR for TCRβ expression with CD3 colabeling demonstrated which of these lesions was a B-cell-rich T-cell lymphoma. We conclude that the RT in situ PCR methodology will allow the routine determination of monoclonal vs multiclonal expression patterns of the TCRβ using archival paraffin-embedded tissues.