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Single–Cell Light Scatter as a Probe of Refractile Body Formation in Recombinant Escherichia Coli

Bio/Technology volume 6, pages 423–426 (1988)Cite this article

Abstract

Accumulation of foreign proteins as inclusion bodies in Escherichia coli was investigated using single–cell light scatter measurements from flow cytometry. Significant increases in forward–angle light scatter and right–angle light scatter were observed for E. coli strains overproducing two types of interferon and two types of veterinary growth hormone. The original host morphology before the induction of foreign gene expression was found to have a significant effect on the subsequent enhancement of light scatter by inclusion body formation. The light scatter distributions obtained by flow cytometry differed significantly from volume distributions obtained using Coulter sizing. Thus flow cytometry may provide information on cloned gene product accumulation at the single–cell level that is not available from other methods.

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References

  1. Shively, J.M. 1974. Inclusion bodies of prokaryotes. Ann. Rev. of Microbiol. 28:167–187.

    Article  CAS  Google Scholar 

  2. Schoemaker, J., Clark, J., Saukkonen, J. 1981. Characterization of polysaccharide accumulations in a cell division defective mutant of Escherichia coli 15T−. J. of Gen. Microb. 123:323–333.

    CAS  Google Scholar 

  3. Prouty, W., Goldberg, A. 1972. Fate of abnormal proteins in E. coli: Accumulation in intracellular granules before catabolism. Nature 240:147–150.

    Article  CAS  Google Scholar 

  4. Paul, D., Van Frank, R., Muth, W., Ross, J., Williams, D. 1983. Immunocytochemical demonstration of human proinsulin chimeric polypeptide within cytoplasmic inclusion bodies of Escherichia coli. Eur. J. of Cell Biol. 31:171–174.

    CAS  Google Scholar 

  5. Williams, D., Van Frank, R.M., Muth, W., Burnett, J. 1982. Cytoplasmic inclusion bodies in Escherichia coli producing biosynthetic human insulin proteins. Science 215:687–689.

    Article  CAS  Google Scholar 

  6. Schoemaker, J., Brasnett, A., Marston, F. 1985. Examination of calf prochymosin accumulation in Escherichia coli: Disulphide linkages are a structural component of prochymosin-containing inclusion bodies EMBO J 4:775–786.

    Article  CAS  Google Scholar 

  7. Fieschko, J., Ritch, T., Bengston, D., Fenton, D., Mann, M. 1985. The relationship between cell dry weight concentration and culture turbidity for a recombinant E. coli K12 strain producing high levels of human alpha interferon analogue. Biotechnology Progress 1:205–208.

    Article  CAS  Google Scholar 

  8. Schoner, R.G., Lee, E.F., Schoner, B.E. 1985. The isolation and purification of protein granules from E. coli Cells overproducing bovine growth hormone. Bio/Technology 3:151–154.

    Article  CAS  Google Scholar 

  9. Chen, Y. 1983. Increased buoyant densities of protein overproducing Escherichia coli Cells. Biochem. & Biophys. Res. Comm. 111:104–111.

    Article  Google Scholar 

  10. Von Meyenberg, K., Jorgensen, B., Van Deurs, B. 1984. Physiological and morphological effects of overproduction of membrane-bound ATP synthase in Escherichia coli K-12. EMBO J. 3:1791–1797.

    Article  Google Scholar 

  11. Georgiou, G., Telford, J., Shuler, M., Wilson, B. 1986. Localization of inclusion bodies in Escherichia coli overproducing β-lactamase or alkaline phosphatase. Appl. & Env. Microb. 52:1157–1161.

    CAS  Google Scholar 

  12. Taylor, G., Hoare, M., Gray, D., Marston, F. 1986. Size and density of protein inclusion bodies. Bio/Technology 4:553–557.

    CAS  Google Scholar 

  13. Mas, J., Pedros-Alio, C., Guerrero, R. 1985. Mathematical model for determining the effects of intracytoplasmic inclusions on volume and density of microorganisms. J. Bacteriology 164:749–756.

    CAS  Google Scholar 

  14. Mullaney, P., Dean, P. 1970. The small angle light scattering of biological Cells. Biophysical J. 10:764–772.

    Article  CAS  Google Scholar 

  15. Koch, A. 1968. Theory of the angular dependence of light scattered by bacteria and similar-sized biological objects. J. Theoret. Biology 18:133–156.

    Article  CAS  Google Scholar 

  16. Wyatt, P. 1968. Differential light scattering. A physical method for identifying living bacterial Cells. Applied Optics 7:1879–1896.

    Article  CAS  Google Scholar 

  17. Latimer, P., Moore, D., Bryant, F. 1968. Changes in total light scattering and absorption caused by changes in particle conformation. J. Theoret. Biol. 21:348–367.

    Article  CAS  Google Scholar 

  18. Schafer, I. 1979. Multiangle light scattering flow photometry of cultured human fibroblasts: Comparison of normal Cells with a mutant line containing cytoplasmic inclusions. J. of Histochem & Cytochem. 27:359–365.

    Article  CAS  Google Scholar 

  19. Srienc, F., Arnold, B., Bailey, J.E. 1984. Characterization of intracellular accumulation of poly-β-hydroxybutyrate (PHB) in individual cells of Alcaligenes eutrophus H16 by flow cytometry. Biotech. & Bioeng. 26:982–987.

    Article  CAS  Google Scholar 

  20. Fiel, R., Munson, B. 1970. Small angle scattering of bioparticles II. Cells and Cellular organelles. Exp. Cell Res. 59:421–428.

    Article  CAS  Google Scholar 

  21. Normark, S. 1969. XXX J. Bact. 98:1274–1280.

    CAS  PubMed  Google Scholar 

  22. Shapiro, H. 1985. Practical Flow Cytometry, p. 88. Alan R. Liss, Inc., New York.

    Google Scholar 

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Author notes

  1. M. B. Mann, D. M. Fenton and L. B. Tsai: Amgen Inc. 1900 Oak Terrace Lane, Thousand Oaks, California 91320.

Authors and Affiliations

  1. Department of Chemical Engineering, California Institute of Technology, Pasadena, California, 91125

    K. D. Wittrup, M. B. Mann, D. M. Fenton, L. B. Tsai & J. E. Bailey

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  1. K. D. Wittrup
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  2. M. B. Mann
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  3. D. M. Fenton
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  4. L. B. Tsai
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  5. J. E. Bailey
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Wittrup, K., Mann, M., Fenton, D. et al. Single–Cell Light Scatter as a Probe of Refractile Body Formation in Recombinant Escherichia Coli. Nat Biotechnol 6, 423–426 (1988). https://doi.org/10.1038/nbt0488-423

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