How I Diagnose Mature T-Cell Proliferations by Flow Cytometry

Steven H. Kroft, MD; Alexandra M. Harrington, MD


Am J Clin Pathol. 2022;158(4):456-471. 

In This Article

Abstract and Introduction


Objectives: Mature T-cell neoplasms are a challenging area of diagnostic hematopathology. Flow cytometry has emerged as a useful technique for T-cell assessment.

Methods: We discuss the application of flow cytometry to the evaluation of mature T-cell proliferations, to include illustrative cases, theoretical framework, detailed review of normal and reactive T-cell subsets, and examination of diagnostic pitfalls.

Results: Immunophenotypic aberrancy can be construed as a direct expression of the neoplastic phenotype, in contrast to clonal expansion, which is seen in reactive and neoplastic T-cell proliferations. Major and minor T-cell subsets show characteristic patterns of antigen expression. Reactive states can manifest expansions of normal minor subsets and also show alterations of antigen expression on certain populations. However, some patterns of antigen expression are either never or very rarely encountered in reactive T cells. Flow cytometric tools are now available to directly assess clonality in specific T-cell populations. Technical and biological pitfalls may complicate the interpretation of T-cell flow cytometry.

Conclusions: Flow cytometry is a very useful tool in the diagnostic armamentarium for the assessment of mature T-cell proliferations, but it must be interpreted based on a thorough knowledge of the T-cell immune response, as well as an awareness of clinical context.


The accurate interpretation of mature T-cell proliferations has historically been challenging for diagnostic pathologists for a number of reasons. Because of their relative rarity, most Western pathologists have limited experience with them. Robust classification of peripheral T-cell lymphomas (PTCLs) lagged significantly behind that of B-cell non-Hodgkin lymphomas, and as a consequence, detailed investigation lagged as well. At a histologic level, PTCLs frequently are composed of complex mixtures of cell types, and there is often morphologic overlap with reactive proliferations. Consequently, effective and reliable diagnostic tools are needed for accurate and confident diagnosis.

In recent years, flow cytometry has emerged as a useful method for investigating mature T-cell proliferations. While not without its challenges, due to both technical and biologic issues, in most situations it can provide a clear-cut assessment of the nature of various T-cell processes. This review begins with two illustrative cases, followed by a general discussion of the application of flow cytometry to the interpretation of mature T-cell proliferations. Two specific challenging diagnostic areas are then discussed, and the article ends with a discussion of the salient features of the two illustrative cases.