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Comparison of LR white resin, lowicryl K4M and Epon postembedding procedures for immunogold staining of actin in the testis

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Summary

The efficiency of various postembedding procedures for actin immunogold detection was compared using testicular tissue as a model. Whatever the fixative, testes embedded in LR White resin or in Lowicryl K4M showed few differences as regard ultrastructural preservation and gave similar actin antigenicity preservation. A purified polyclonal antibody (IgG) and a monoclonal antibody (IgM) visualized with gold secondary antibody yielded high labeling intensity whereas the IgG-protein-A gold association was less efficient. Crude antisera gave a low specific staining/background ratio. Samples of testes, fixed in different conditions, were also embedded in Epon, omitting propylene oxide and lowering polymerization temperature to 40°–50° C. This slight modification improved ultrastructural preservation which was better than with hydrophilic resins, as well as made possible immunogold detection of actin though antigenicity preservation was lesser than with these resins. Thus, in Epon embedded samples actin labeling, using IgG antiactin-gold secondary antibody, was similar to that observed after hydrophilic resin-protein-A gold procedures. In addition to actin labeling of various somatic cells it was confirmed that actin is a consistent component of the subacrosomal space of spermatids during the greater part of spermiogenesis in rat.

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References

  • Altmann LG, Schneider BG, Papermaster DS (1984) Rapid embedding of tissues in Lowicryl K4M for immunoelectron microscopy. J Histochem Cytochem 32:1217–1223

    Google Scholar 

  • Bendayan M (1984) Protein A-gold electron microscopic immunocytochemistry: methods, applications, and limitations. J Electron Microsc Tech 1:243–270

    Google Scholar 

  • Bendayan M, Nanci A, Kan FWK (1987) Effect of tissue processing on colloidal gold cytochemistry. J Histochem Cytochem 35:983–996

    Google Scholar 

  • Camatini M, Anelli G, Casale A (1986) Identification of actin in boar spermatids and spermatozoa by immunoelectron microscopy. Eur J Cell Biol 42:311–318

    Google Scholar 

  • Camatini M, Casale A, Cifarelli M (1988) Immunocytochemical identification of actin in rabbit spermiogenesis and spermatozoa. Eur J Cell Biol 45:274–281

    Google Scholar 

  • Carlemalm E, Garavito RM, Villiger W (1982) Resin development for electron microscopy and an analysis of embedding at low temperature. J Microsc (London) 126:123–143

    Google Scholar 

  • Forssmann WG, Ito S, Weihe E, Aoki A, Dym M, Fawcett DW (1977) An improved perfusion fixation method for the testis. Anat Rec 188:307–314

    Google Scholar 

  • Fouquet JP, Kann ML, Dadoune JP (1989) Immunogold distribution of actin during spermiogenesis in rat, hamster, monkey and man. Anat Rec 223 (in press)

  • Franke WW, Grund C, Fink A, Weber K, Jockusch BM, Zentgraf H, Osborn M (1978) Location of actin in the microfilaments bundles associated with the junctional specializations between Sertoli cells and spermatids. Biol Cell 31:7–14

    Google Scholar 

  • Gosselin JE, Cate CC, Pettengill OS, Sorenson GD (1986) Immunocytochemistry: its evolution and criteria for its application in the study of Epon-embedded cells and tissue. Am J Anat 175:135–160

    Google Scholar 

  • Gounon P, Karsenti E (1981) Involvement of contractile proteins in the changes in consistency of oocyte nucleoplasm of the newt Pleurodeles waltlii. J Cell Biol 88:410–421

    Google Scholar 

  • Halenda RM, Primakoff P, Myles DG (1987) Actin filaments, localized to the region of the developing acrosome during early stages, are lost during later stages of guinea pig spermiogenesis. Biol Reprod 36:491–499

    Google Scholar 

  • Hisano S, Adachi T, Macgawa M, Daikoku S (1986) Some improvement in tissue preparation and colloidal-gold immunolabeling for electron microscopy. Am J Anat 175:245–266

    Google Scholar 

  • Kellenberger E, Durrenberger M, Villerger W, Carlemalm E, Wurtz M (1987) The efficiency of immunolabel on Lowicryl sections compared to theoretical predictions. J Histochem Cytochem 35:959–969

    Google Scholar 

  • Loesser EK, Doane KJ, Wilson FJ, Roisen FJ, Malamed S (1986) Improved immunoelectron microscopic method for localizing cytoskeletal proteins in Lowicryl K4M embedded tissues. J Histochem Cytochem 34:1477–1485

    Google Scholar 

  • Mar H, Tsukada T, Gown AM, Wight, TN, Baskin DG (1987) Correlative light and electron microscopic immunocytochemistry on the same section with colloidal gold. J Histochem Cytochem 35:419–425

    Google Scholar 

  • Masri BA, Russell LD, Vogl AW (1987) Distribution of actin in spermatids and adjacent Sertoli cell regions of the rat. Anat Rec 218:20–26

    Google Scholar 

  • Newman GR, Hobot JA (1987) Modern acrylics for post-embedding immunostaining techniques. J Histochem Cytochem 35:971–981

    Google Scholar 

  • Russell LD, Weber JE, Vogl AW (1986) Characterization of filaments within the subacrosomal space of rat spermatids during spermiogenesis. Tissue Cell 18:887–898

    Google Scholar 

  • Russell LD, Vogl AW, Weber JE (1987a) Actin localization in male germ cell intercellular bridges in the rat and ground squirrel and disruption of bridges by cytochalasin D. Am J Anat 180:25–40

    Google Scholar 

  • Russell LD, Amlani SR, Vogl AW, Weber JE (1987b), Characterization of filaments within Leydig cells of the rat testis. Am J Anat 178:231–240

    Google Scholar 

  • Timms BG (1986) Postembedding immunogold labeling for electron microscopy using “LR White” resin. Am J Anat 175:267–275

    Google Scholar 

  • Toyama Y (1986) Actin-like filaments in the Sertoli cell junctional specializations in the swine and mouse testis. Anat Rec 186:477–492

    Google Scholar 

  • Toyama Y (1977) Actin-like filaments in the myoid cell of the testis. Cell Tissue Res 177:221–226

    Google Scholar 

  • Vogl AW, Soucy LJ, Lew GJ (1985) Distribution of actin in isolated seminiferous epithelia and denuded tubule walls of the rats. Anat Rec 213:63–71

    Google Scholar 

  • Vogl AW, Grove BD, Lew GJ (1986) Distribution of actin in Sertoli cell ectoplasmic specializations and associated spermatids in the ground squirrel testis. Anat Rec 215:331–341

    Google Scholar 

  • Welch JE, O'Rand MG (1985) Identification and distribution of actin in spermatogenic cells and spermatozoa of the rabbit. Dev Biol 109:411–417

    Google Scholar 

  • Wolosewick J, De Mey J (1982) Localization of tubulin and actin in polyethylene glycol embedded rat seminiferous epithelium. Biol Cell 44:85–88

    Google Scholar 

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Authors and Affiliations

  1. Groupe d'Etude de la Formation et de la Maturation du Gamète Mâle, Laboratoire d'Histologie, UER Biomédicale, 45 rue des Saints-Pères, F-75270, Paris Cedex 06, France

    M. -L. Kann & J. -P. Fouquet

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  1. M. -L. Kann
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  2. J. -P. Fouquet
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Kann, M.L., Fouquet, J.P. Comparison of LR white resin, lowicryl K4M and Epon postembedding procedures for immunogold staining of actin in the testis. Histochemistry 91, 221–226 (1989). https://doi.org/10.1007/BF00490136

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  • DOI: https://doi.org/10.1007/BF00490136

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