Detection of double-negative T cells and gating strategy. Peripheral blood lymphocytes were analyzed by flow cytometry for DNTs (CD4^-/CD8^-, CD3⁺, TCRαβ⁺). A histogram of CD3-FITC versus side scatter (A,C) was used to gate on T cells (CD3⁺/low SS). The events in the T-cell gate were sent to second histogram (B,D) of CD4 and CD8-RD1 versus TCRαβ-PC5. Double-negative T cells are depicted in lower right quadrant of panels B and D. Panel B represents a patient with a normal test (1.2% DNTs), and panel D represents a patient with an elevated test (8.9% DNTs). SS LIN indicates side scatter linear.