Monitoring of NeoR⁺ persistently circulating GMCs. (A) Kinetics of persistently circulating NeoR⁺ GMCs in 4 patients with a follow-up of more than 10 years. △ and ● indicate the absolute number of lymphocytes and GMCs, respectively (right y-axis); □ indicates the percentage of GMCs among circulating lymphocytes (assuming that there is one transgene copy of per cell) (left y-axis). The last analysis date was day 3557 (118.5 months), 3228 (118.5 months), 3379 (112.6 months), and 2063 (68.76 months) for patients 6, 7, 8, and 9, respectively. (B) Absence of T-cell clonality. Clonality was determined by PCR analysis of TCRγ gene rearrangements within donor GMCs before infusion, within PBMCs, and within GMCs G418-reselected at different time points after BMT, using 2 Vγ-Jγ PCR mixes (PCR A-TCRg: V γ 1f + V γ 10 − J γ 1.1/2.1 + J γ 1.3/2.3; PCR B-TCRg: V γ 9 + V γ 11 − J γ 1.1/2.1 + J γ 1.3/2.3). Both PCRs used consensus-specific primers from Vγ families, allowing coverage of most of the TCRγ rearrangements (sensitivity, 10⁻² to 10⁻³). The Gaussian profile confirms the absence of detectable clonal expansion. Multiple peaks without Gaussian distribution were in favor of an oligoclonal T-cell population. Polyclonal and monoclonal controls were DNA extracted from tonsil and Jurkat cell lines, respectively. N/A indicates not analyzable.