Flow cytometric assessment of peripheral blood contamination and proliferative activity of human bone marrow cell populations

Abstract

Bone marrow aspiration is superior to bone marrow biopsies due to less discomfort to the volunteer or patient, but it is inferior concerning the reproducibility of cytokinetic information. Therefore, a method that could select aspirates of quality and reproducibility equal to those of biopsies was sought. Low‐density (mononucleated) bone marrow cells were labelled with T200 common leukocyte antigen, CD45, which differentiate cells into erythroid, myeloid, and lymphocyte + monocyte subpopulations based on their immunofluorescence intensity. A hypotonic propidium iodide solution was added, and DNA cell cycle characteristics of the total cells and the subpopulations were obtained. Twenty‐two aspirations were performed on three healthy men. There was a strong negative correlation between the amount of CD45‐gated lymphocytes + monocytes, indicative of peripheral blood cell contamination in the aspirate, and the percentage of total cells and subpopulations in DNA S phase. A marked reduction in the percentage of cells in S phase was observed when the lymphocyte + monocyte counts were higher than 30%; this level was used to exclude aspirates with an unacceptable degree of peripheral blood cell admixture. Twelve of the aspirates were found to be of acceptable quality due to their low lymphocyte + monocyte count. These aspirates were compared with 11 bone marrow biopsy expellates from hematologically normal patients undergoing open cardiac surgery. The 12 aspirates were found to have almost identical mean percent S‐phase cells as the biopsy expellates, both for the total cell population (14% ± 3.45% vs. 15% ± 1.5%) and for the erythroid (24% ± 6% vs. 24.4% ± 3.3%) and myeloid (10% ± 2.4% vs. 10.7% ± 2.5%) subpopulations. The aspirates with an unacceptable degree of peripheral blood cell admixture had a significantly lower mean percent cells in S phase for both the total cells and the subpopulations. It is concluded that bone marrow aspirates of small volumes may give cytokinetic information as reproducible as bone marrow biopsies if the above‐described method is applied.