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. 1974 Jan 1;60(1):8–25. doi: 10.1083/jcb.60.1.8

CYTOCHEMISTRY OF GOLGI FRACTIONS PREPARED FROM RAT LIVER

Marilyn G Farquhar 1, J J M Bergeron 1, George E Palade 1
PMCID: PMC2109143  PMID: 4358430

Abstract

Cytochemical tests for several marker enzymes were applied to liver tissue and to the three Golgi fractions (GF1, GF2, GF3) separated by the procedure of Ehrenreich et al. from liver homogenates of alcohol-treated rats. 5'-Nucleotidase (AMPase) reaction product was found in all three fractions but in different locations: It occurred along the inside of the membrane of VLDL-filled vacuoles in GF1 and GF2, and along the outside of the cisternal membranes in GF3. In the latter it was restricted to the dilated cisternal rims and was absent from the cisternal centers. The AMPase activity found in the fractions by biochemical assay is therefore indigenous to Golgi components and is not due to contamination by plasma membrane. Acid phosphatase (AcPase) reaction product was detected within lysosomal contaminants in GF1 and within many VLDL-filled vacuoles in GF1 and GF2, indicating that AcPase activity is due not only to contaminating lysosomes, but also to enzyme indigenous to Golgi secretory vacuoles. G-6-Pase reaction product was present in GF3 and within contaminating endoplasmic reticulum fragments, but not in other fractions. Thiamine pyrophosphatase (TPPase) was localized to some of the VLDL-filled vacuoles and cisternae in GF1 and GF2, and was not found in the cisternae in GF3. The results demonstrate the usefulness of cytochemical methods in monitoring the fractionation procedure: They have (a) allowed a reliable identification of contaminants, (b) made possible a distinction between indigenous and contaminating activities, and (c) shown, primarily by the results of the TPPase test, that the procedure achieves a meaningful subfractionation of Golgi elements, with GF1 and GF3, representing primarily trans-Golgi elements from the secretory Golgi face, and GF3 consisting largely of cis-Golgi components from the opposite face.

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Selected References

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  1. Bainton D. F., Farquhar M. G. Differences in enzyme content of azurophil and specific granules of polymorphonuclear leukocytes. II. Cytochemistry and electron microscopy of bone marrow cells. J Cell Biol. 1968 Nov;39(2):299–317. doi: 10.1083/jcb.39.2.299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bainton D. F., Farquhar M. G. Segregation and packaging of granule enzymes in eosinophilic leukocytes. J Cell Biol. 1970 Apr;45(1):54–73. doi: 10.1083/jcb.45.1.54. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bergeron J. J., Ehrenreich J. H., Siekevitz P., Palade G. E. Golgi fractions prepared from rat liver homogenates. II. Biochemical characterization. J Cell Biol. 1973 Oct;59(1):73–88. doi: 10.1083/jcb.59.1.73. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cheetham R. D., Morré D. J., Pannek C., Friend D. S. Isolation of a Golgi apparatus-rich fraction from rat liver. IV. Thiamine pyrophosphatase. J Cell Biol. 1971 Jun;49(3):899–905. doi: 10.1083/jcb.49.3.899. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Claude A. Growth and differentiation of cytoplasmic membranes in the course of lipoprotein granule synthesis in the hepatic cell. I. Elaboration of elements of the Golgi complex. J Cell Biol. 1970 Dec;47(3):745–766. doi: 10.1083/jcb.47.3.745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. DiPietro D. L., Zengerle F. S. Separation and properties of three acid phosphatases from human placenta. J Biol Chem. 1967 Jul 25;242(14):3391–3395. [PubMed] [Google Scholar]
  7. Farquhar M. G., Bainton D. F., Baggiolini M., de Duve C. Cytochemical localization of acid phosphatase activity in granule fractions from rabbit polymorphonuclear leukocytes. J Cell Biol. 1972 Jul;54(1):141–156. doi: 10.1083/jcb.54.1.141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Farquhar M. G., Palade G. E. Cell junctions in amphibian skin. J Cell Biol. 1965 Jul;26(1):263–291. doi: 10.1083/jcb.26.1.263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Friend D. S. Cytochemical staining of multivesicular body and golgi vesicles. J Cell Biol. 1969 Apr;41(1):269–279. doi: 10.1083/jcb.41.1.269. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Friend D. S., Farquhar M. G. Functions of coated vesicles during protein absorption in the rat vas deferens. J Cell Biol. 1967 Nov;35(2):357–376. doi: 10.1083/jcb.35.2.357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. GOLDFISCHER S., ESSNER E., NOVIKOFF A. B. THE LOCALIZATION OF PHOSPHATASE ACTIVITIES AT THE LEVEL OF ULTRASTRUCTURE. J Histochem Cytochem. 1964 Feb;12:72–95. doi: 10.1177/12.2.72. [DOI] [PubMed] [Google Scholar]
  12. Goldfischer S., Essner E., Schiller B. Nucleoside diphosphatase and thiamine pyrophosphatase activities in the endoplasmic reticulum and golgi apparatus. J Histochem Cytochem. 1971 Jun;19(6):349–360. doi: 10.1177/19.6.349. [DOI] [PubMed] [Google Scholar]
  13. Hamilton R. L., Regen D. M., Gray M. E., LeQuire V. S. Lipid transport in liver. I. Electron microscopic identification of very low density lipoproteins in perfused rat liver. Lab Invest. 1967 Feb;16(2):305–319. [PubMed] [Google Scholar]
  14. Heinrikson R. L. Purification and characterization of a low molecular weight acid phosphatase from bovine liver. J Biol Chem. 1969 Jan 25;244(2):299–307. [PubMed] [Google Scholar]
  15. NOVIKOFF A. B., GOLDFISCHER S. Nucleosidediphosphatase activity in the Golgi apparatus and its usefulness for cytological studies. Proc Natl Acad Sci U S A. 1961 Jun 15;47:802–810. doi: 10.1073/pnas.47.6.802. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Nichols B. A., Bainton D. F., Farquhar M. G. Differentiation of monocytes. Origin, nature, and fate of their azurophil granules. J Cell Biol. 1971 Aug;50(2):498–515. doi: 10.1083/jcb.50.2.498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Novikoff P. M., Novikoff A. B., Quintana N., Hauw J. J. Golgi apparatus, GERL, and lysosomes of neurons in rat dorsal root ganglia, studied by thick section and thin section cytochemistry. J Cell Biol. 1971 Sep;50(3):859–886. doi: 10.1083/jcb.50.3.859. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nyquist S. E., Mollenhauer H. H. A Golgi apparatus acid phosphatase. Biochim Biophys Acta. 1973 Jul 5;315(1):103–112. doi: 10.1016/0005-2744(73)90134-4. [DOI] [PubMed] [Google Scholar]
  19. REYNOLDS E. S. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol. 1963 Apr;17:208–212. doi: 10.1083/jcb.17.1.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Smith J. K., Whitby L. G. The heterogeneity of prostatic acid phosphatase. Biochim Biophys Acta. 1968 Mar 25;151(3):607–618. doi: 10.1016/0005-2744(68)90007-7. [DOI] [PubMed] [Google Scholar]
  21. Touster O., Aronson N. N., Jr, Dulaney J. T., Hendrickson H. Isolation of rat liver plasma membranes. Use of nucleotide pyrophosphatase and phosphodiesterase I as marker enzymes. J Cell Biol. 1970 Dec;47(3):604–618. doi: 10.1083/jcb.47.3.604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Widnell C. C. Cytochemical localization of 5'-nucleotidase in subcellular fractions isolated from rat liver. I. The origin of 5'-nucleotidase activity in microsomes. J Cell Biol. 1972 Mar;52(3):542–558. doi: 10.1083/jcb.52.3.542. [DOI] [PMC free article] [PubMed] [Google Scholar]

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