Combined negative and positive selection of mobilized CD34⁺ blood cells

We tested four negative and two positive selection methods for separation of CD34⁺ cells from mobilized blood cells, and analysed fold‐enrichment, purity and recovery of CD34⁺ cells after selection procedures. The elimination of mature CD34⁻ cells was achieved by adhesion to nylon‐wool fibre (5.9 ± 1.0 mean fold‐enrichment and 65.2 ± 2.3 mean recovery of CD34⁺ cells). Standard or modified Ficoll‐Hypaque and Percoll density gradients, as well as phagocytosis with magnetic beads, were less effective in eliminating CD34⁻ cells, both purity and fold‐enrichment of CD34⁺ cells being lower than those obtained with separation by nylon‐wool. Both positive selection methods tested, Ceprate and MiniMacs System, generated highly purified CD34⁺ cell populations ranging from 80% to 90%. The recovery of CD34⁺ cells was optimal with MiniMacs (77.9±3.6) and low with Ceprate (28.8±2.8). Based on these results, in two large‐scale experiments we combined nylon‐wool fibre and MiniMacs System in a two‐step separation procedure obtaining a 36.9±2.6 mean fold‐enrichment and a 50.5±0.3 mean recovery of CD34⁺ cells. In this way we achieved optimal enrichment and recovery of CD34⁺ cells, with a substantial saving of cost compared to either selection method alone.