Fig. 2. Dual staining for T-cell receptor constant β chain (TRBC)1 and TRBC2 demonstrates mutually exclusive and polytypic TRBC expression on benign T-cells and T-cell receptor variable β (TCR-Vβ) subsets.

A Representative peripheral blood flow cytometry findings on a healthy donor showing polytypic TRBC expression on total, CD4⁺ (cyan) and CD8⁺ (orange) T-cells. B TRBC2:TRBC1 ratios of total, CD4⁺ and CD8⁺ T-cells from 25 healthy donor’s peripheral blood specimens. Solid lines: medians. Dotted lines: thresholds for clonality. C TCR-Vβ repertoire by flow cytometry on gated TRBC1⁺ (black bars) and TRBC1^- (gray bars) peripheral blood T-cells from five healthy donors, showing remarkably similar distributions. *p < 0.05. D TRBC2:TRBC1 ratios of each TCR-Vβ-positive T-cell subset from five healthy donors. Dotted lines depict thresholds for clonality. E Representative peripheral blood flow cytometry plots from a healthy donor showing distinct naïve, central memory (T_CM), effector memory (T_EM), and effector memory with reacquired CD45RA (T_EMRA) subsets on the CD4⁺ (cyan) and CD8⁺ (orange) T-cell compartments. F TRBC2:TRBC1 ratios of gated naïve-memory CD4⁺ and CD8⁺ T-cell subsets from 10 healthy donors. Solid lines: medians. Dotted lines: thresholds for clonality.