Abstract
A new assay for the precise measurement of microbial killing by leukocytes is presented. The method assumes that release of radioactively labeled DNA from the microbe is direct evidence of cell death. Human peripheral blood leukocytes incubated with [14C]thymidine-labeled Salmonella typhimurium released 32 to 59% of the radioactivity after 4 h and 63 to 75% after 18 h. Inactivated leukocytes released less than 5% of the radioactivity. None of the released radioactivity is retained within the leukocyte, and 60% remains precipitable with trichloroacetic acid. Leukocytes released substantial radioactivity from labeled Escherichia coli but only a slight amount from staphylococci. Mouse peritoneal macrophages were also shown to release radioactivity from Salmonella. The DNA release assay avoids the errors inherent in prior killing methods which measure viability by growth inhibition. It is rapid, reproducible, and highly specific.
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Selected References
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- Ambrose E. J., Antia N. H., Khonolkar S. R. Uptake of radioactive DOPA by M. leprae. Nature. 1974 Jun 28;249(460):854–855. doi: 10.1038/249854a0. [DOI] [PubMed] [Google Scholar]
- BRAUDE A. I., FELTES J. Studies in the destruction of staphylococci by human leukocytes; effect of clumping of intracellular and extracellular bacteria on the results obtained with the agar-plate method. J Lab Clin Med. 1953 Aug;42(2):289–298. [PubMed] [Google Scholar]
- COHN Z. A., MORSE S. I. Interactions between rabbit polymorphonuclear leucocytes and staphylococci. J Exp Med. 1959 Sep 1;110:419–443. doi: 10.1084/jem.110.3.419. [DOI] [PMC free article] [PubMed] [Google Scholar]
- COHN Z. A. The fate of bacteria within phagocytic cells. I. The degradation of isotopically labeled bacteria by polymorphonuclear leucocytes and macrophages. J Exp Med. 1963 Jan 1;117:27–42. doi: 10.1084/jem.117.1.27. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Carr H. S., Rosenkranz H. S. H3-thymidine and the conservation of deoxyribonucleic acid. J Bacteriol. 1966 Dec;92(6):1840–1841. doi: 10.1128/jb.92.6.1840-1841.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cline M. J. A new white cell test which measures individual phagocyte function in a mixed leukocyte population. I. A neutrophil defect in acute myelocytic leukemia. J Lab Clin Med. 1973 Feb;81(2):311–316. [PubMed] [Google Scholar]
- Cole P., Brostoff J. Intracellular killing of Listeria monocytogenes by activated macrophages (Mackaness system) is due to antibiotic. Nature. 1975 Aug 7;256(5517):515–517. doi: 10.1038/256515a0. [DOI] [PubMed] [Google Scholar]
- Elsbach P., Beckerdite S., Pettis P., Franson R. Persistence of regulation of macromolecular synthesis by Escherichia coli during killing by disrupted rabbit granulocytes. Infect Immun. 1974 Apr;9(4):663–668. doi: 10.1128/iai.9.4.663-668.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Elsbach P., Pettis P., Beckerdite S., Franson R. Effects of phagocytosis by rabbit granulocytes on macromolecular synthesis and degradation in different species of bacteria. J Bacteriol. 1973 Aug;115(2):490–497. doi: 10.1128/jb.115.2.490-497.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
- FLECK D. G. Mouse protection and enhancement of phagocytosis by antisera to Streptococcus pyogenes. Br J Exp Pathol. 1956 Aug;37(4):406–414. [PMC free article] [PubMed] [Google Scholar]
- Feingold D. S., Goldman J. N., Kuritz H. M. Locus of the lethal event in the serum bactericidal reaction. J Bacteriol. 1968 Dec;96(6):2127–2131. doi: 10.1128/jb.96.6.2127-2131.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Friedlander A. M. DNA release as a direct measure of microbial killing. I. Serum bactericidal activity. J Immunol. 1975 Nov;115(5):1404–1408. [PubMed] [Google Scholar]
- Ghuysen J. M. Use of bacteriolytic enzymes in determination of wall structure and their role in cell metabolism. Bacteriol Rev. 1968 Dec;32(4 Pt 2):425–464. [PMC free article] [PubMed] [Google Scholar]
- HILL G. A., MARCUS S. Study of cellular mechanisms in resistance to systemic Histoplasma capsulatum infection. J Immunol. 1960 Jul;85:6–13. [PubMed] [Google Scholar]
- HIRSCH J. G., STRAUSS B. STUDIES ON HEAT-LABILE OPSONIN IN RABBIT SERUM. J Immunol. 1964 Jan;92:145–154. [PubMed] [Google Scholar]
- Jones T. C., Hirsch J. G. The interaction in vitro of Mycoplasma pulmonis with mouse peritoneal macrophages and L-cells. J Exp Med. 1971 Feb 1;133(2):231–259. doi: 10.1084/jem.133.2.231. [DOI] [PMC free article] [PubMed] [Google Scholar]
- STAHELIN H., KARNOVSKY M. L., SUTER E. Studies on the interaction between phagocytes and tubercle bacilli. II. The action of phagocytes upon C14-labelled tubercle bacilli. J Exp Med. 1956 Jul 1;104(1):137–150. doi: 10.1084/jem.104.1.137. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tan J. S., Watanakunakorn C., Phair J. P. A modified assay of neutrophil function: use of lysostaphin to differentiate defective phagocytosis from impaired intracellular killing. J Lab Clin Med. 1971 Aug;78(2):316–322. [PubMed] [Google Scholar]
- Taylor-Robinson D., Purcell R. H., Wong D. C., Chanock R. M. A colour test for the measurement of antibody to certain mycoplasma species based upon the inhibition of acid production. J Hyg (Lond) 1966 Mar;64(1):91–104. doi: 10.1017/s0022172400040377. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Veale D. R., Smith H., Witt K. Penetration of penicillin into human phagocytes containing gonococci. Lancet. 1975 Feb 8;1(7902):306–308. doi: 10.1016/s0140-6736(75)91210-6. [DOI] [PubMed] [Google Scholar]
- WILSON A. T., WILEY G. G., BRUNO P. Fate of non-virulent group A streptococci phagocytized by human and mouse neutrophils. J Exp Med. 1957 Dec 1;106(6):777–786. doi: 10.1084/jem.106.6.777. [DOI] [PMC free article] [PubMed] [Google Scholar]
