Detection of the JAK2 V617F mutation in FTOC-derived T-cell subpopulations from PV and IMF CD34⁺ cells. CD34⁺ cells (50 000) were cultured in fetal thymic lobes for 3 weeks as indicated in “Patients, materials, and methods.” At the end of the culture, cells collected from each thymic lobe were labeled by FITC-conjugated anti–human CD45, PE anti–human CD4, and APC anti–human CD8 monoclonal antibodies and analyzed by flow cytometry. Human CD45⁺ CD4⁺CD8⁻, CD4⁻CD8⁺, CD4⁻CD8⁻, and CD4⁺CD8⁺ fractions were subsequently sorted and sequenced. The immunophenotypic scattergrams and sequence traces of initial CD34⁺ cells and FTOC T-cell fractions from patients IMF6 (A) and IMF7 (B) are shown. The red arrow indicates the presence of a small mutant T peak at the threshold of detection.