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The Separation of Cytotoxic Human Peripheral Blood Monocytes Into High and Low Phagocytic Subsets by Centrifugal Elutriation
Author(s) -
Chiu K.M.,
McPherson L.H.,
Harris J.E.,
Braun D.P.
Publication year - 1984
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1002/jlb.36.6.729
Subject(s) - elutriation , monocyte , cytotoxicity , cytotoxic t cell , microbiology and biotechnology , biology , peripheral blood mononuclear cell , chromatography , flow cytometry , collagenase , immunology , chemistry , biochemistry , in vitro , enzyme , organic chemistry
Mononuclear cells (4 × 10 8 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 51 Cr‐labeled Chang liver cell line. Similarly, each mono‐ cyte‐containing fraction was found to mediate the same level of cytotoxicity against antibody‐sensitized 51 Cr‐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.