The Separation of Cytotoxic Human Peripheral Blood Monocytes Into High and Low Phagocytic Subsets by Centrifugal Elutriation

Abstract Mononuclear cells (4 × 108 in a 5-ml volume) were loaded onto a Beckman JE-6B elutriator rotor spinning at 2,000 ± 10 rpm with a flow rate of 10.0 ml/ min. After lymphocytes were removed at flow rates between 10 and 11 ml/min with 500 ml of buffer, the flow rate was increased by 0.5 ml/min/fraction to collect 100-ml fractions. Highly enriched monocytes as judged by nonspecific esterase staining and morphology (70–95%) were found in each of eight fractions collected with flow rates between 11.5 to 15.0 ml/min. When stimulated with phorbol myristic acetate, these fractions mediated equivalent levels of cytotoxicity against 51Cr-labeled Chang liver cell line. Similarly, each mono- cyte-containing fraction was found to mediate the same level of cytotoxicity against antibody-sensitized 51Cr-labeled Chang liver cells. In contrast, cytotoxicity against the natural killer cell-sensitive K-562 cell line was found in only those fractions that contained a high percentage of lymphocytes. The fractions that were enriched in monocytes were found to differ in their ability to ingest latex. Those monocyte fractions that were collected between 11.0–12.0 ml/min consisted primarily of low numbers of latex-ingesting monocytes (<30%). Those monocyte fractions that were collected between 12.5 and 15.0 ml/min consisted primarily of latex-ingesting monocytes (50–70%). These data show that cytotoxic monocytes can be separated by centrifugal elutriation into at least two subsets that can be distinguished by their phagocytic activity.