Summary
Thein situ identification of lymphocyte subpopulations by means of immunopathological techniques using specific monoclonal antibodies provides a tool for the study of the gastrointestinal-associated lymphoid tissue (GALT) in health and disease. In this field, monoclonal antibodies have been applied previously using light microscopy and either immunofluorescence or immunoperoxidase; however, these techniques are not sensitive enough to allow precise evaluation of localization of labelling. We describe an immunoelectronmicroscopic method, which defines labelling specificity, since it allows the identification of cells by immunophenotype labelling and ultrastructural markers simultaneously. This in turn allows a better evaluation of the labelled cells and of the relationship between labelled and unlabelled cells.
The main features of the method are the use of fresh tissue samples, fixing in paraformaldehyde CaCl2, and the coupling of the immune reaction to an amplification system (avidin-biotin-peroxidase complex). The technique yields a good preservation of cellular ultrastructure, together with a strong and specific immunolabelling.
Our results confirm the high specificity of monoclonal antibodies when applied to immunopathology techniques. We confirm the pattern of distribution of various lymphocyte subsets in the jejunal mucosa described by other authors by light microscopy.
Similar content being viewed by others
References
Abo T, Balch CM (1981) A differentiation antigen of human NK and K cells identified by a monoclonal antibody. (HNK-1) J Immunol 127: 1024–1029
Berti E, Monti M, Cavicchini S, Caputo R (1983) The avidin-biotin peroxidase complex (ABC Px), in situ immunoelectron microscopy. Arch Dermatol Res 275:134–138
Bland PW, Richens ER, Britton DC, Lloyd JV (1979) Isolation and purification of human large bowel mucosal lymphoid cells: effect of separation technique on functional characteristic. Gut 20:1037–1046
Chiba M, Bartnik W, Re Mine SH, Thayer WR, Shorter RG (1981) Human colonic intraepithelial and lamina propria lymphocytes: cytotoxicy in vitro and the popential effects of the isolation methods on their functional properties. Gut 22:177–186
Colucci G, Colombo M, Del Ninno E, Paronetto F (1983) In situ characterization by monoclonal antibodies of the mononuclear infiltrate in chronic active hepatitis. Gastroenterology 85:1138–1145
Engleman EG, Warnke R, Fox RI, Levy R (1981) Studies of a human T lymphocytes antigen recognized by a monoclonal antibody. Proc Natl Acad Sci USA 78:1791–1795
Engleman EG, Benike CJ, Glickman E, Evans RL (1982) Antibodies to membrane structures that distinguish Suppressor-Cytotoxic and Helper T lymphocyte subpopulations block the mixed leukocyte reaction in man. J Exp Med 154:193–198
Ferguson A (1977) Intraepthelial lymphocytes of the small intestine. Gut 18:921–938
Ferguson A, Mowat A, McI (1980) Immunological mechanisms in the small intestine. In: Wright R (ed) Recent advances in gastrointestinal pathology. WB Saunders, London, pp 255–266
Ferguson A (1983 a) Why study T-cell subsets in Crohn's diseases? Gut 24:687–691
Ferguson A, Strobel S (1983 b) Immunology and physiology of digestion. In: Lessof MH (ed) Clinical reactions to food. John Wiley and Sons, Chichester, pp 59–86
Gibson PR, Verhaar HJJ, Selby WS, Jewell DP (1984) The mononuclear cells of human mesenteric blood intestinal mucosa and mesenteric lymphonodes: compartimentalization of NK cells. Clin Exp Immunol 56:445–452
Hanau D, Fabre M, Stampf JL, Grosshaus E (1983) Langerhans cells in mouse epidermia (letter). J Invest Dermatol 81:290
Janossy G, Tidman N, Papageorgion ES, Kung PC, Goldstein G (1981) Distribution of T lymphocyte subsets in the human bone marrow and thymus, an analysis with monoclonal antibodies. J Immunol 126 (4):1608–1613
Karnovsky MJ (1965) A formaldehyde-glutaraldehyde fixative of high osmolarity for use in electron microscopy (abstr). J Cell Biol 27:137a
Ljunghall K, Loof L, Forsum U (1982) T lymphocyte subsets in the duodenal epithelium in Dermatitis Herpetiformis. Acta Dermatol (Stockholm) 62:485–489
Martin PJ, Hansen JA, Siadak AW, Nowinski RC (1981) Monoclonal antibodies recognizing human T lymphocytes and malignant human B lymphocytes: a comparative study. J Immunol 127:1920–1923
McClean JW, Nakane PK (1974) Periodate-lysine-paraformaldehyde fixative, a nex fixative for immunoelectron microscopy. J Histochem Cytochem 22:1077–1083
Mowat A, McI, Ferguson A (1981) Hypersensitivity reactions in the small intestine. 5 Induction of cell mediated immunity to a dietary antigen. Clin Exp Immunol 43:574–582
Reinherz EL, Scholssman SF (1980) The differentiation and function of human T lymphocytes. Cell 19:821–827
Selby WS, Janossy G, Goldstein G, Jewell DP (1981 a) T lymphocyte subsets in human intestinal mucosa: the distribution and relationship to MHC-derived antigens. Clin Exp Immunol 44:453–458
Selby WS, Janossy G, Jewell DP (1981 b) Immunological characterization of intraepithelial lymphocytes of the human gastrointestinal tract. Gut 22:169–176
Selby WS, Janossy G, Bofill M, Jewell DP (1983) Lymphocyte subpopulation in the human small intestine. The findings in normal mucosa and in the mucosa of patients with adult coeliac disease. Clin Exp Immunol 52:219–228
Selby WS, Janossy G, Bofill M, Jewell DP (1984) Intestinal lymphocyte subpopulation in inflammatory bowel disease: an analysis by immunohistological and cell isolation techniques. Gut 25:32–40
Taylor CR, Hofman FM, Modlin RL, Rea TH (1983) Immunoperoxidase techniques applied to dermatopathology. J Cutan Pathol 38:145–163
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Vecchi, M., Berti, E., Primignani, M. et al. In situ identification of immune competent cells in gastrointestinal mucosa: An evaluation by immunoelectronmicroscopy. Vichows Archiv A Pathol Anat 406, 407–415 (1985). https://doi.org/10.1007/BF00710232
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00710232