TRBC1/CD3 dual color immunohistochemistry: An effective test to support a diagnosis of T‐cell lymphoma in cytology specimens

The diagnosis of T-cell lymphoma is challenging, and establishing clonality is an important aspect of the workup. Molecular testing to establish T-cell clonality though T-cell receptor (TCR) gene rearrangement polymerase chain reaction is the most common modality and can detect clonal rearrangements in ∼90% of T-cell lymphomas, but is limited by increased turnaround time and cost and the need for sufficient material for testing. In the maturation of T cells, the TCR locus rearrangement will result in the mutually exclusive expression of either the TCR beta-chain constant domain 1 or 2 (TRBC1, TRBC2). Antibodies to TRBC1 and TRBC2 have been used in the setting of flow cytometry, and more recently immunohistochemistry, to establish T cell clonality. This study evaluates the utility of a TRBC1/CD3 dual stain for the diagnosis of T-cell lymphoma. The laboratory information system was searched from 2019 through 2025 for cytology samples with a diagnosis of T-cell lymphoma. Twelve samples were identified along with controls. Dual color immunohistochemistry for TRBC1 and CD3 was performed using a brown chromogen for TRBC1 and a red chromogen for CD3. The proportion of cells staining for TRBC1 and CD3 was scored. A monoclonal T-cell population was detected in all samples of T-cell lymphoma. All B-cell lymphomas and benign samples showed a polyclonal staining pattern consisting of a mixture of brown and red cells. Using immunohistochemistry for TRBC1 paired with CD3 is a reliable surrogate for T-cell clonality testing in cytology specimens and can support a diagnosis of T-cell lymphoma on limited material.