Summary
Experiments were conducted to determine the capacity of various enzyme preparations to dissociate single cells from guinea pig lung tissue. The number of cells separated from tissue progressively increased as the concentration of crude trypsin was increased from 25 to 250 mg per 100 ml. This action could be inhibited by soy bean trypsin inhibitor. Elastase, but not ethylenediaminetetraacetate (disodium salt), crystalline trypsin, nor chymotrypsin, dissociated cells from lung tissues. Crude trypsin (Trypsin 1∶300) was found to contain 3.0 Sachar units of elastase per mg. Elastase was also inhibited by soy bean trypsin inhibitor. Only some collagenase preparations dissociated cells from lung tissue. Impure bacterial proteases dissociated lung cells. Our data suggest that the term “trypsinization” to denote dissociation of cells from tissue with crude preparations of trypsin is misleading and should be discontinued.
Similar content being viewed by others
References
Uriel, J., and M. Kaminski. 1961. Héterogénéité des préparations de trypsine etudieés par electrophorèse en gélose. C. R. Acad. Sci. (Paris) 253: 573–575.
Pine, L., G. C. Taylor, D. M. Miller, G. Bradley, and H. R. Wetmore. 1969. Comparison of good and bad lots of trypsin used in the production of primary monkey kidney cells. A definition of the problem and comparison of certain enzymatic characteristics. Cytobios 1: 197–207.
Shotton, D. M. 1970. Elastase. In: G. E. Perlmann and L. Lorand (Eds.),Methods in Enzymology. Vol. 19. Academic Press, New York, pp. 113–140.
Moscona, A. A. 1963. Inhibition by trypsin inhibitors of dissociation of embryonic tissue by trypsin. Nature (London) 199: 379–380.
Essner, E., H. Sato, and M. Belkin. 1954. Experiments on ascites hepatoma. I. Enzymatic digestion and alkaline degradation of the cementing substance and separation of cells in tumor islands. Exp. Cell Res. 7: 430–437.
Marcus, P. I., S. J. Cieciura, and T. T. Puck. 1956. Clonal growth in vitro of epithelial cells from normal human tissues. J. Exp. Med. 104: 615–628.
Edwards, A. L. 1964.Statistical Analysis. Holt, Rinehart and Winston, New York, p. 130.
Bergmeyer, H. 1965.Methods of Enzymatic Analysis. Academic Press, New York p. 999.
Walford, R. L., and B. Kickhoffen. 1962. Selective inhibition of elastolytic and proteolytic properties of elastase. Arch. Biochem. Biophys. 98: 191–196.
Gertler, A., and Y. Birk. 1970. Isolation and characterization of porcine proelastase. Eur. J. Biochem. 12: 170–176.
Schubert, M., and D. Hamerman. 1968.A Primer on Connective Tissue Biochemistry. Lea and Febiger. Philadelphia, p. 46.
Pumper, R. W., P. Fagan, and W. G. Taylor. 1971. Trypsin sensitivity of mammalian cells grown in a serum-free medium. In Vitro 6: 266–268.
Hinz, R. W. and J. T. Syverton. 1959. Mammalian cell cultures for study of influenza virus. I. Preparation of monolayer cultures with collagenase. Proc. Soc. Exp. Biol. Med. 101: 19–22.
Rinaldini, L. M. 1959. An improved method for the isolation and quantitative cultivation of embryonic cells. Exp. Cell Res. 16:477–505.
Author information
Authors and Affiliations
Additional information
Partially supported bv Armour-Baldwin Laboratories and the National Institute of Health, Grant, AM 12919.
Rights and permissions
About this article
Cite this article
Phillips, H.J. Dissociation of single cells from lung or kidney tissue with elastase. In Vitro 8, 101–105 (1972). https://doi.org/10.1007/BF02615967
Issue Date:
DOI: https://doi.org/10.1007/BF02615967