Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1968 Sep 1;38(3):615–627. doi: 10.1083/jcb.38.3.615

ULTRASTRUCTURE OF HUMAN LEUKOCYTES AFTER SIMULTANEOUS FIXATION WITH GLUTARALDEHYDE AND OSMIUM TETROXIDE AND "POSTFIXATION" IN URANYL ACETATE

James G Hirsch 1, Martha E Fedorko 1
PMCID: PMC2108377  PMID: 4874495

Abstract

Human leukocytes in suspension or in monolayer cultures have been processed for electron microscopy by fixation in a freshly made cold mixture of glutaraldehyde and osmium tetroxide and by "postfixation" in uranyl acetate. Simultaneous exposure to glutaraldehyde and osmium tetroxide eliminates many of the shortcomings seen when either of these agents is used alone as the initial fixative. Specimens are processed to the stage of dehydration as single cell suspensions or as very small clumps to assure rapid penetration of fixatives and efficient washing. The technique is rapid and reproducible. Electron micrographs presented in this report illustrate the ultrastructural features of human white cells prepared by this method.

Full Text

The Full Text of this article is available as a PDF (1.3 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. ACKERMAN G. A., GRASSO J. A., KNOUFF R. A. Morphological and histochemical studies of the leukemic cells from a patient with atypical myeloblastic leukemia with special reference to intracytoplasmic mucopolysaccharide vacuoles and fibrillar formation. Blood. 1960 Sep;16:1253–1267. [PubMed] [Google Scholar]
  2. Anderson D. R. Ultrastructure of normal and leukemic leukocytes in human peripheral blood. J Ultrastruct Res. 1966 Sep;9:1–42. doi: 10.1016/s0022-5320(66)90002-5. [DOI] [PubMed] [Google Scholar]
  3. BESSIS M., THIERY J. P. Electron microscopy of human white blood cells and their stem cells. Int Rev Cytol. 1961;12:199–241. doi: 10.1016/s0074-7696(08)60541-0. [DOI] [PubMed] [Google Scholar]
  4. Bainton D. F., Farquhar M. G. Origin of granules in polymorphonuclear leukocytes. Two types derived from opposite faces of the Golgi complex in developing granulocytes. J Cell Biol. 1966 Feb;28(2):277–301. doi: 10.1083/jcb.28.2.277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. DE PETRIS S., KARLSBAD G., PERNIS B. Filamentous structures in the cytoplasm of normal mononuclear phagocytes. J Ultrastruct Res. 1962 Aug;7:39–55. doi: 10.1016/s0022-5320(62)80025-2. [DOI] [PubMed] [Google Scholar]
  6. De Duve C., Wattiaux R. Functions of lysosomes. Annu Rev Physiol. 1966;28:435–492. doi: 10.1146/annurev.ph.28.030166.002251. [DOI] [PubMed] [Google Scholar]
  7. FLOREY H. W., GRANT L. H. Leucocyte migration from small blood vessels stimulated with ultraviolet light: an electron-microscope study. J Pathol Bacteriol. 1961 Jul;82:13–17. doi: 10.1002/path.1700820103. [DOI] [PubMed] [Google Scholar]
  8. FREEMAN J. A., SAMUELS M. S. The ultrastructure of a fibrillar formation of leukemic human blood. Blood. 1958 Aug;13(8):725–731. [PubMed] [Google Scholar]
  9. Fedorko M. E., Hirsch J. G., Cohn Z. A. Autophagic vacuoles produced in vitro. I. Studies on cultured macrophages exposed to chloroquine. J Cell Biol. 1968 Aug;38(2):377–391. doi: 10.1083/jcb.38.2.377. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. HIRSCH J. G., CHURCH A. B. Studies of phagocytosis of group A streptococci by polymorphonuclear leucocytes in vitro. J Exp Med. 1960 Mar 1;111:309–322. doi: 10.1084/jem.111.3.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hirsch J. G., Fedorko M. E., Cohn Z. A. Vesicle fusion and formation at the surface of pinocytic vacuoles in macrophages. J Cell Biol. 1968 Sep;38(3):629–632. doi: 10.1083/jcb.38.3.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. KELLENBERGER E., RYTER A., SECHAUD J. Electron microscope study of DNA-containing plasms. II. Vegetative and mature phage DNA as compared with normal bacterial nucleoids in different physiological states. J Biophys Biochem Cytol. 1958 Nov 25;4(6):671–678. doi: 10.1083/jcb.4.6.671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. LOCKWOOD W. R., ALLISON F. ELECTRON MICROGRAPHIC STUDIES OF PHAGOCYTIC CELLS. I. MORPHOLOGICAL CHANGES OF THE CYTOPLASM AND GRANULES OF RABBIT GRANULOCYTES ASSOCIATED WITH INGESTION OF ROUGH PNEUMOCOCCUS. Br J Exp Pathol. 1963 Dec;44:593–600. [PMC free article] [PubMed] [Google Scholar]
  14. LUFT J. H. Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol. 1961 Feb;9:409–414. doi: 10.1083/jcb.9.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. MERCER E. H. An electron microscopic study of Amoeba proteus. Proc R Soc Lond B Biol Sci. 1959 Mar 17;150(939):216–232. doi: 10.1098/rspb.1959.0016. [DOI] [PubMed] [Google Scholar]
  16. NOVIKOFF A. B., ESSNER E., QUINTANA N. GOLGI APPARATUS AND LYSOSOMES. Fed Proc. 1964 Sep-Oct;23:1010–1022. [PubMed] [Google Scholar]
  17. RABINOWITZ Y. SEPARATION OF LYMPHOCYTES, POLYMORPHONUCLEAR LEUKOCYTES AND MONOCYTES ON GLASS COLUMNS, INCLUDING TISSUE CULTURE OBSERVATIONS. Blood. 1964 Jun;23:811–828. [PubMed] [Google Scholar]
  18. Sutton J. S., Weiss L. Transformation of monocytes in tissue culture into macrophages, epithelioid cells, and multinucleated giant cells. An electron microscope study. J Cell Biol. 1966 Feb;28(2):303–332. doi: 10.1083/jcb.28.2.303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. TANAKA Y. FIBRILLAR STRUCTURES IN THE CELLS OF BLOODFORMING ORGANS. J Natl Cancer Inst. 1964 Sep;33:467–485. [PubMed] [Google Scholar]
  20. Trump B. F., Bulger R. E. New ultrastructural characteristics of cells fixed in a glutaraldehyde-osmium tetroxide mixture. Lab Invest. 1966 Jan;15(1 Pt 2):368–379. [PubMed] [Google Scholar]
  21. VENABLE J. H., COGGESHALL R. A SIMPLIFIED LEAD CITRATE STAIN FOR USE IN ELECTRON MICROSCOPY. J Cell Biol. 1965 May;25:407–408. doi: 10.1083/jcb.25.2.407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Watanabe I., Donahue S., Hoggatt N. Method for electron microscopic studies of circulating human leukocytes and observations on their fine structure. J Ultrastruct Res. 1967 Oct 31;20(5):366–382. doi: 10.1016/s0022-5320(67)80106-0. [DOI] [PubMed] [Google Scholar]
  23. Wetzel B. K., Horn R. G., Spicer S. S. Fine structural studies on the development of heterophil, eosinophil, and basophil granulocytes in rabbits. Lab Invest. 1967 Mar;16(3):349–382. [PubMed] [Google Scholar]
  24. ZUCKER-FRANKLIN D., HIRSCH J. G. ELECTRON MICROSCOPE STUDIES ON THE DEGRANULATION OF RABBIT PERITONEAL LEUKOCYTES DURING PHAGOCYTOSIS. J Exp Med. 1964 Oct 1;120:569–576. doi: 10.1084/jem.120.4.569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. ZUCKER-FRANKLIN D. THE ULTRASTRUCTURE OF CELLS IN HUMAN THORACIC DUCT LYMPH. J Ultrastruct Res. 1963 Oct;59:325–339. doi: 10.1016/s0022-5320(63)80010-6. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES

OSZAR »