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Method for enrichment of proliferating myeloid cells from normal and leukemic human bone marrow.

A method is described for the enrichment of immature myeloid cells from normal human bone marrow. The procedure consists of three consecutive steps: density gradient centrifugation, hypotonic shock, and velocity sedimentation. The results of 10 isolations are reported in this paper. The unprocessed bone marrow contained an average of 8.9% immature myeloid cells. Cell differential counts of interface fractions, obtained after density gradient (Flcoll-lsopaque 1.068 g/ml) centrifugation, revealed an average of 27.2% immature myeloid cells. Subsequently, erythroid precursors were removed by hypotonic shock. The average concentration of immature myeloid cells increased to 36.4%. After the final step, velocity sedimentation at 1 g, fractions were collected with an average of 62.6% immature myeloid cells, of which 25.4% were in S-phase. Bone marrow cells from five patients with full-blown (>85% blast cells) acute myeloid leukemia were subjected to the same procedure, excluding hypotonic shock. The mean percentage of S-phase cells in the unprocessed bone marrow was 9.7%. Interface fractions (FicollIsopaque 1.074 g/ml) consisted of more than 90% leukemic myeloblasts, of which 9.8% (mean value) were in S-phase. After velocity sedimentation a separation was obtained in fractions with mean percentages of DNA-synthesizing cells ranging from 3.8% to 26.6%.