Summary
The use of an immunohistochemical method permits the localization of creatine kinase isoenzymes MM and BB in tissue sections. Frozen sections are first incubated with the specific antiserum and secondly with the soluble antigen under investigation. The antibody fixed creatine kinase can then be visualized by the tetrazolium-salt linked histochemical reaction.
In this way CK-BB was found in the smooth muscle and the mucosa of the human colon. In sections of skeletal muscle CK-MM was predominantly localized in the intermyofibrillar space. Membrane bound activity could be demonstrated in the sarcoplasmic reticulum and the surface membrane after elution of the cytoplasmic enzyme.
In the human tonsilla CK-BB was localized in lymphatic and epithelial tissues, CK-MM in the muscle fibers. The isoenzyme patterns in single sections of tonsilla were in parallel determined by the immunotitration assay. The results indicate the usefulness of the combined application of histochemistry and immunotitration in serial tissue sections.
Zusammenfassung
In Gefrierschnitten lassen sich die Isoenzyme der Creatinkinase MM und BB durch spezifische Antikörper selektiv präzipitieren. Nach Sättigung der freien Antikörpervalenzen durch Zugabe von exogenem Antigen wird das im Schnitt fixierte Isoenzym durch eine histochemische Technik lokalisiert. Die Methode ermöglicht eine isoenzymspezifische Histochemie der Creatinkinase.
Die Anwendung dieser Technik am menschlichen Gewebe führte zu folgenden Ergebnissen: CK-BB ließ sich sowohl in der Muskulatur als auch in der Mucosa des Colons nachweisen, während im Skelettmuskel zwischen cytoplasmatischer CK-MM und membrangebundenem Enzym im sarkoplasmatischen Reticulum und im Sarkolemm differenziert werdn konnte. In der Tonsille wurden CK-BB in epithelialem Gewebe, CK-MM in Muskelfasern lokalisiert. Das Isoenzymmuster von benachbarten einzelnen Schnitten wurde parallel immunologisch analysiert. Auf den Vorteil der Kombination von Immuntitration und Histchemie an gleichen Schnitten wird verwiesen.
Similar content being viewed by others
Literatur
Baba, N., Kim, S., Farrell, E.C.: Histochemistry of Creatine Phosphokinase. J. mol. cell. Cardiol. 8, 559–617 (1976)
Baskin, R.J., Deamer, D.W.: A membrane-bound creatine phosphokinase in fragmented sarcoplasmic reticulum. J. biol. Chem. 245, 1345–1347 (1970)
Dawson, D.M., Eppenberger, H.M., Kaplan, N.O.: Creatine kinase: Evidence for dimeric structure. Biochem. biophys. Res. Commun. 21, 346–353 (1965)
Eppenberger, H.M., Walliman, T., Kuhn, H.J., Turner, D.C.: Localization of creatine kinase isozymes in muscle cells: physiological significance. In: Isozymes II 409–423 (1975)
Fitch, C.D., Schields, R.P.: Creatine metabolism in skeletal muscle. I. Creatine movement across muscle membranes. J. biol. Chem. 241, 3611–3614 (1966)
Jacobs, H., Heldt, H.W., Klingenberg, M.: High activity of creatine kinase in mitochondria from muscle and brain and evidence for a separate mitochondrial enzyme of creatine kinase. Biochem. biophys. Res. Commun. 6, 516–521 (1964)
Jocker-Wretou, E., Pfleiderer, G.: Quantitation of creatine kinase isoenzymes in human tissue and sera by an immunological method. Clin. Chim. Acta 58, 223–232 (1975)
Khan, M.A., Holt, P.G., Knight, J.O., Kakulas, B.A.: Incubation film technique for the histochemica localization of creatine kinase. Histochemie 26, 120–125 (1971)
Khan, M.A., Holt, P.G., Papadimitriou, J.R., Knight, J.O., Kakulas, B.A.: Creatine kinase, a histochemical study by the gelatine film lead precipitation technique. Histochemie 32, 49–58 (1972)
Khan, M.A., Papadimitriou, J.M., Holt, P.G., Kakulas, B.A.: Distribution of creatine kinase in mammalian nervous tissue: A cytochemical study. In: Basic research in myology. Proc. 2nd int. Cong. Muscle Diseases, Perth, pp. 66–71. Amsterdam: Excerpta medica 1973
Lammers, J.M.J., Hülsmann, W.C.: The effect of fructose on the stores of energy rich phosphate in rat jejunum in vivo. Biochim. biophys. Acta 313, 1–8 (1973)
Pfeiderer, G., Thöner, M., Wachsmuth, E.D.: Histological examination of the aldolase monomer composition of cells from human kidney and hypernephroid carcinoma. Beitr. Path. 156, 226–279 (1975)
Robin, Y., Benyamin, Y., van Thoai, N.: Existance of homologous antigenic structures in unfolded creatine kinase and arginine kinase. Febs Lett. 63, 174–178 (1976)
Saks, V.A., Lipina, N.V., Shalov, V.G., Smirnow, V.N., Chazov, E., Grosse, R.: The localization of the MM isozyme of creatine phosphokinase on the surface membrane of myocardial cells and its functional coupling to ouabain-inhibited Na+/K+-ATPase. Biochim. biophys. Acta 465, 550 (1977)
Sherwin, A.C., Karpati, G., Bulcke, J.A.: Immunohistochemical localization of creatine phosphokinase in skeletal muscle. Proc. Nat. Acad. Sci. 64, 171–175 (1969)
Shields, R.P., Whitehair, C.K., Carrow, H.E., Heusner, W.W., van Huss, W.D.: Skeletal muscle function and structure after depletion of creatine. Lab. Invest. 33, 151–158 (1975)
Sjövall, K.: A tetrazolium technique for the histochemical localization of ATP: creatine phosphotransferase. Histochemie 10, 336–340 (1967)
Tsung, S.H.: Creatine kinase isoenzyme pattern in human tissue obtained at surgery. Clin. Chem. 22, 173–175 (1976)
Wachsmuth, E.D., Thöner, M., Pfleiderer, G.: The cellular distribution of aldolase isoenzymes in rat kidney and brain determined in tissue sections by the immuno-histochemical method. Histochemistry 45, 143–161 (1975)
Wegmann, R., Kishino, Y., El Sammanoudy, F.A.: Histochemical study on creatine kinase of developing and denervated rat muscle as compared with glycolytic and oxidative enzymes. Ann. Histochem. 19, 227–238 (1974)
Author information
Authors and Affiliations
Additional information
Unterstützt durch das Schwerpunktprogramm „Enzymdiagnostik” Pf 32/36 der Deutschen Forschungsgemeinschaft, 5300 Bonn-Bad Godesberg
Rights and permissions
About this article
Cite this article
Jockers-Wretou, E., Giebel, W. & Pfleiderer, G. Immunohistochemische Lokalisierung der Isoenzyme der Creatinkinase im menschlichen Gewebe. Histochemistry 54, 83–95 (1977). https://doi.org/10.1007/BF00493332
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00493332