Detection of HTLV-1–specific CD4⁺ T cells by in vitro activation with Env and Tax peptides.

(A) Dot plots showing intracellular cytokine staining for IFN-γ production in CD4⁺ T cells from a patient with HAM/TSP (TAF) and an uninfected control (UN). PBMCs were depleted of CD8⁺ T cells, cultivated with Env and Tax peptides for 6 hours in vitro, then harvested and stained. One representative experiment from 3 patients with HAM/TSP is shown. Numbers in brackets show actual numbers of events acquired. (B) Comparison of HTLV-1–specific CD4⁺ T-cell frequencies determined by independent Elispot assays at different time points from a single blood sample. One representative experiment from 3 patients with HAM/TSP is shown. IFN-γ SFCs were divided by the number of CD4⁺ T cells present in each well, then multiplied by 100 to provide the data shown. A response was defined as positive if the number of spots exceeded the mean + 2 SDs of the spot count in the negative control wells (no peptides added). (C) Comparison of HTLV-1–specific CD4⁺ T-cell frequencies determined by Elispot and intracellular cytokine staining from the same blood sample. Results from 3 patients with HAM/TSP (TAT, TAF, TW) are shown. IFN-γ SFCs are divided by the number of CD4⁺ T cells present in each well, then multiplied by 100 to provide the Elispot data shown. Flow cytometric data shown is derived from IFN-γ⁺, CD4⁺ T cells divided by the total number of CD4⁺ T cells present, then multiplied by 100.