Telomere length of input CB CD34⁺ cells and suspension cells obtained at weekly intervals in long-term OP9/adeno-TPO stromal cell coculture and during stroma-free LTC. (A) TRF Southern blot analyses at baseline and weekly. (B) Mean or peak TRF values plotted over 18 weeks. Cultured cells showed some initial telomere elongation as measured by mean TRF values with a loss of less than 10% of initial value (1 kb). Peak TRF showed no decline, suggesting that the population was heterogeneous with a substantial subpopulation showing no telomere loss over the entire culture period. Telomere length of input CB CD34⁺ cells and of CD34⁺ cells purified from cytokine-stimulated, nonstromal cultures at intervals over 20 weeks of culture (C) and after repeated expansion-isolation procedures over 20 weeks of culture (D). Telomere length has been determined by flow FISH. Note the initial increase in telomere length over the first 8 weeks (C-D), the progressive decline beginning at 12 weeks (C) and the stabilization of the telomere size over 16 weeks and the progressive decline beginning at 20 weeks (D).