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. 1984 Mar 1;218(2):441–447. doi: 10.1042/bj2180441

Preparation of rat enterocyte mitochondria.

B Masola, D F Evered
PMCID: PMC1153358  PMID: 6424661

Abstract

Rat enterocyte mitochondria were prepared with respiratory control ratios of 4 or 5 and occasionally 6. When EGTA was excluded from the mitochondrial incubation medium the calculated P/O ratios were high, especially those based on the first addition of ADP. These ratios were lowered by increasing the EGTA concentration from 1 mM to 2 mM in the mitochondrial preparation medium and including 1 mM-EGTA in the incubation medium. The use of EDTA in the enterocyte isolation medium led to the mitochondria requiring added cytochrome c. Substituting EGTA for EDTA abolished this requirement. The mitochondrial fraction consisted of two components, an upper cream-coloured layer rich in DNA and a lower brown-coloured layer poor in DNA. Both components were capable of oxidative phosphorylation with succinate or the glutamate/malate couple as substrates. The mitochondrial yield was assessed by assaying succinate dehydrogenase activity, and the contamination of the mitochondrial fraction by other cell organelles was assessed by assays for appropriate marker enzymes.

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Selected References

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

  1. BURTON K. A study of the conditions and mechanism of the diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem J. 1956 Feb;62(2):315–323. doi: 10.1042/bj0620315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. CHANCE B., WILLIAMS G. R. The respiratory chain and oxidative phosphorylation. Adv Enzymol Relat Subj Biochem. 1956;17:65–134. doi: 10.1002/9780470122624.ch2. [DOI] [PubMed] [Google Scholar]
  3. Chappell J. B. The oxidation of citrate, isocitrate and cis-aconitate by isolated mitochondria. Biochem J. 1964 Feb;90(2):225–237. doi: 10.1042/bj0900225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Evans E. M., Wrigglesworth J. M., Burdett K., Pover W. F. Studies on epithelial cells isolated from guinea pig small intestine. J Cell Biol. 1971 Nov;51(21):452–464. doi: 10.1083/jcb.51.2.452. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. HARRER D. S., STERN B. K., REILLY R. W. REMOVAL AND DISSOCIATION OF EPITHELIAL CELLS FROM THE RODENT GASTROINTESTINAL TRACT. Nature. 1964 Jul 18;203:319–320. doi: 10.1038/203319a0. [DOI] [PubMed] [Google Scholar]
  6. HUEBSCHER G., WEST G. R. SPECIFIC ASSAYS OF SOME PHOSPHATASES IN SUBCELLULAR FRACTIONS OF SMALL INTESTINAL MUCOSA. Nature. 1965 Feb 20;205:799–800. doi: 10.1038/205799a0. [DOI] [PubMed] [Google Scholar]
  7. Harrison D. D., Webster H. L. The preparation of isolated intestinal crypt cells. Exp Cell Res. 1969 May;55(2):257–260. doi: 10.1016/0014-4827(69)90489-3. [DOI] [PubMed] [Google Scholar]
  8. Huang K. C. Uptake of L-tyrosine and 3-0-methylglucose by isolated intestinal epithelial cells. Life Sci. 1965 Jun;4(12):1201–1206. doi: 10.1016/0024-3205(65)90333-4. [DOI] [PubMed] [Google Scholar]
  9. Iemhoff W. G., van den Berg J. W., de Pijper A. M., Hülsmann W. C. Metabolic aspects of isolated cells from rat small intestinal epithelium. Biochim Biophys Acta. 1970 Aug 14;215(2):229–241. doi: 10.1016/0304-4165(70)90020-6. [DOI] [PubMed] [Google Scholar]
  10. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  11. NAKAMURA M., PICHETTE P., BROITMAN S., BEZMAN A. L., ZAMCHECK N., VITALE J. J. Studies on the inhibitory effect of intestinal mucosa of succinoxidase activity. J Biol Chem. 1959 Jan;234(1):206–210. [PubMed] [Google Scholar]
  12. Perris A. D. Isolation of the epithelial cells of the rat small intestine. Can J Biochem. 1966 Jun;44(6):687–693. doi: 10.1139/o66-086. [DOI] [PubMed] [Google Scholar]
  13. SHERRATT H. S., HUBSCHER G. Properties of mitochondrial preparations from the small-intestinal mucosa of the guinea-pig. Biochim Biophys Acta. 1963 Feb 5;69:403–405. doi: 10.1016/0006-3002(63)91274-5. [DOI] [PubMed] [Google Scholar]
  14. STANBURY P. J. Comparison of the mitochondria of the small intestine of vertebrates. Nature. 1961 Oct 7;192:67–67. doi: 10.1038/192067a0. [DOI] [PubMed] [Google Scholar]
  15. Stern B. K., Jensen W. E. Active transport of glucose by suspensions of isolated rat intestinal epithelial cells. Nature. 1966 Feb 19;209(5025):789–790. doi: 10.1038/209789a0. [DOI] [PubMed] [Google Scholar]
  16. Watford M., Lund P., Krebs H. A. Isolation and metabolic characteristics of rat and chicken enterocytes. Biochem J. 1979 Mar 15;178(3):589–596. doi: 10.1042/bj1780589. [DOI] [PMC free article] [PubMed] [Google Scholar]

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