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Ultrastructural immunogold labelling of human langerhans cells enriched epidermal cell suspension

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Summary

Colloidal gold particles are well suited as markers in electron microscopy. Indirect immunogold staining was used to identify cell membrane antigens defined by monoclonal antibodies OKT6 and BL6 on human Langerhans cells (LC) in suspensions. Isolated epidermal cells were obtained by skin trypsinization and enriched or depleted in OKT6 positive on BL6 positive LC using the panning method: incubation of OKT6 or BL6 preincubated cells on immunoglobulin coated dishes. Indirect immunogold staining was then performed after prefixation in 2% paraformaldehyde. In LC enriched suspensions, only LC exhibited a specific membrane labelling with OKT6 or BL6 recognized by the presence of small evently distributed gold granules. Neither Birbeck granules, nor other cytoplasmic organelles, were labelled. No other epidermal cells were found positive. In LC depleted suspensions, no labelling was observed. Immunogold labelling on LC enriched suspensions after panning is now in progress for the qualitative evaluation and the quantitative analysis of cell surface constituants and antigens expressed by human dendritic epidermal cells.

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References

  1. Chu A, Eisinger M, Soolee J, Takezaki S, Kung P, Edelson RL (1982) Immunoelectronmicroscopic identification of Langerhans cells using a new antigenic marker. J Invest Dermatol 78:177–180

    Google Scholar 

  2. Cohen J, Bouveret E, Pluot M, Revillard JP (1981) Characterization of Langerhans cells using a monoclonal anti HLA-DR (Ia-like) antibody. Arch pathol Lab Med 105:438

    Google Scholar 

  3. Czernielewski J, Faure M, Schmitt D, Thivolet J (1982) In vitro mixed skin cell lymphocyte culture reaction (MSLR) in man: analysis of the epidermal cell and T cell subpopulations. Clin Exp Immunol 50:426–433

    Google Scholar 

  4. Czernielewski J, Schmitt D, Faure M, Thivolet J (1983) Functional and phenotypic analysis of isolated human Langerhans cells and indeterminate cells. Br J Dermatol 108:129–139

    Google Scholar 

  5. De Mey J, Moeremans M, De Waele M, Gevens G, De Brabander M (1982) The IGS (Immunogold staining) method used with monoclonal antibodies. In: Peeters H (ed) Prot. Biol. Fluids. Pergamon Press, Oxford, pp 943–947

    Google Scholar 

  6. De Waele M, De Mey J, Moereman M, Broodtaerts L, Smet L, Van Camp B (1982) Colloidal gold as a marker for the light microscope. Detection of leukocyte cell surface antigens with monoclonal antibodies. J Clin Immunol 2:24S-31S

    Google Scholar 

  7. Eisinger M, Soo Lee JI, Hefton JM, Darzynkiewicz Z, Chiao JW, De Harven E (1979) Human epidermal cell cultures: growth and differentiation in the absence of dermal components or medium supplements. Proc Natl Acad Sci USA 76:5340–5344

    Google Scholar 

  8. Faure M, Eisinger M, Bystryn JC (1981) Pemphigus, pemphigoid and epidermal upper-cytoplasmic antigens: changes in expression in cultured human keratinocytes. Arch Dermatol Res 271:73–82

    Google Scholar 

  9. Fithian E, Kung P, Goldstein G, Rubenfeld M, Fenoglio C, Edelson RL (1981) Reactivity of Langerhans cells with hybridoma antibody. Proc Natl Acad Sci USA 78:2541–2544

    Google Scholar 

  10. Geuze H, Slot J, Van Der Ley P, Schuffer R, Griffith J (1981) Use of colloidal gold particles in double-labelling immunoelectronmicroscopy of ultrathin frozen tissue sections. J Cell Biol 89:653–665

    Google Scholar 

  11. Hannah J, Langley M, Naeim F (1983) Immunogold staining method for identification and subclassification of lymphoid cells and their malignant counterpents. Lab Invest 48: 32A

    Google Scholar 

  12. Harrist TJ, Muhlbauer JE, Murphy GF, Mihm MC Jr, Bahn AK (1983) T6 is superior to Ia (HLA-DR) is a marker for Langerhans cells and indeterminate cells in normal epidermis: A monoclonal antibody study. J Invest Dermatol 80:100–103

    Google Scholar 

  13. Holden CA, Morgan EM, Mac Donald DM (1982) A technique for immuno ultrastructural identification of T6 positive Langerhans cells and indeterminate cells in mycosis fungoides. J Invest Dermatol 79:382–384

    Google Scholar 

  14. Horisberger M (1979) Evaluation of colloidal gold as a cytochemical marker for transmission and scanning electronmicroscopy. Biol Cell 36:253–258

    Google Scholar 

  15. Kung PC, Goldstein G, Reinherz EL, Schlossman SF (1979) Monoclonal antibodies defining distinctive human T cell surface antigens. Science 206:347–349

    Google Scholar 

  16. Lafferty MD, Ackerman GA, Dunn CDR, Lange RD (1981) Ultrastructural, immunocytochemical localization of presumptive erythropoietin binding sites on developing erythrocytic cells of normal human bone marrow. J Histochem Cytochem 29:49–56

    Google Scholar 

  17. Morhenn VB, Benike CJ, Charron DJ, Cox A, Mahrle G, Wood GS, Engelman EG (1982) Use of the fluorescence-activated cell sorter to quantitate and enrich for subpopulations of human skin cells. J Inverst Dermatol 79:277–282

    Google Scholar 

  18. Murphy GF, Bahn AK, Sato S, Harrist TJ, Mihm MD (1981) Characterization of Langerhans cells by the use of monoclonal antibodies. Lab Invest 45:465–468

    Google Scholar 

  19. Murphy GF, Bahn AK, Sato S, Mihm MC, Harrist TJ (1981) A new immunologic marker for human Langerhans cells. N Engl J Med 304:791–792

    Google Scholar 

  20. Page-Faulk W, Taylor GM (1971) An immunocolloid method for the electronmicroscope. Imunochemistry 8:1081–1083

    Google Scholar 

  21. Reinherz EL, Penta AC, Hyssey RE, Schlossman SF (1981) A rapid method for separating functionally intact human T lymphocytes with monoclonal antibodies. Clin Immunol Immunopathol 21:257–266

    Google Scholar 

  22. Roth J, Bendayan M, Orci L (1978) Ultrastructural localization of intracellular antigens by the use of protein A gold complex. J Histochem Cytochem 26:1074–1081

    Google Scholar 

  23. Rowden GR (1980) Expression of Ia antigens on Langerhans cells in mice, guinea pigs and man. J Invest Dermatol 75:22–31

    Google Scholar 

  24. Scheynius A, Klareskog L, Forsum U, Matsson P, Karlsson L, Peterson PA, Sundstrom C (1982) Enrichment of epidermal Langerhans cells. Studies with a monolayer technique and flow cytometry sorting. J Invest Dermatol 82:452–455

    Google Scholar 

  25. Sieber-Blum M, Sieber F, Yamada K (1981) Cellular fibronectin promotes adrenergic differentation of quail neural crest cells in vitro. Exp Cell Res 133:285–295

    Google Scholar 

  26. Wysocki LJ, Sato VL (1978) Panning for lymphocytes: a method for well selection. Proc Natl acad Sci, USA 75:2844–2850

    Google Scholar 

  27. Yonish-Rouach E, Viac J, Schmitt D, Cordier G, Thivolet J, Brochier J (1982) Preparation of monoclonal anti-human thymocyte antibodies: unexpected markers of epidermal cells. In: Serrou B, Rosenfeld C, Daniels J (eds) Current Elsevier North Holland, Biochemical Press, Amsterdam

    Google Scholar 

  28. Yonish-Rouach E, Schmitt D, Brochier J (1983) A monoclonal anti-human thymocyte antibody wich cross reacts with epidermal Langerhans cells. J Immunol Methods (in press)

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Authors and Affiliations

  1. INSERM U 209, Laboratoire de Recherche Dermatologique et Immunologie, Hôpital E. Herriot-Pav. R, F-69374, Lyon Cedex 08, France

    D. Schmitt, M. Faure, C. Dezutter-Dambuyant & D. Tuffery

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  1. D. Schmitt
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  2. M. Faure
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  3. C. Dezutter-Dambuyant
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  4. D. Tuffery
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Schmitt, D., Faure, M., Dezutter-Dambuyant, C. et al. Ultrastructural immunogold labelling of human langerhans cells enriched epidermal cell suspension. Arch Dermatol Res 276, 27–32 (1984). https://doi.org/10.1007/BF00412558

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  • DOI: https://doi.org/10.1007/BF00412558

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