Abstract
The location of the (1→3)-β-glucan, callose, in the walls of pollen tubes in the style of Nicotiana alata Link et Otto was studied using specific monoclonal antibodies. The antibodies were raised against a laminarinhaemocyanin conjugate. One antibody selected for further characterization was specific for (1→3)-β-glucans and showed no binding activity against either a cellopentaose-bovine serum albumin (BSA) conjugate or a (1→3, 1→4)-β-glucan-BSA conjugate. Binding was inhibited by (1→3)-β-oligoglucosides (DP, 3–6) with maximum competition being shown by laminaripentaose and laminarihexaose, indicating that the epitope included at least five (1→3)-β-linked glucopyranose residues. The monoclonal antibody was determined to have an affinity constant for laminarihexaose of 2.7. 104M−1. When used with a second-stage gold-labelled, rabbit anti-mouse antibody, the monoclonal antibody probe specifically located the (1→3)-β-glucan in the inner wall layer of thin sections of the N. alata pollen tubes.
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Abbreviations
- BSA:
-
bovine serum albumin
- PBS:
-
phosphate-buffered saline
- ELISA:
-
enzyme linked immunosorbent assay
- DP:
-
degree of polymerization
- PVC:
-
polyvinyl chloride
References
Anderson, M.A., Harris, P.J., Bonig, I., Clarke, A.E. (1987) Immuno-gold localization of α-l-arabinofuranosyl residues in pollen tubes of Nicotiana alata Link et Otto. Planta 171, 438–442
Dubois, M., Gilles, K.A., Hamilton, J.K., Rebers, P.A., Smith, F. (1956) Colorimetric method for determination of sugars and related substances. Anal. Chem. 28, 350–356
Friguet, B., Chaffotte, A.F., Djavadi-Ohaniance, L., Goldberg, M.E. (1985) Measurement of the true affinity constant in solution of antigen-antibody complexes by enzyme-linked immunosorbent assay. J. Immunol. Methods 77, 305–319
Galfré, G., Howe, S.C., Milstein, C., Butcher, C.W., Howard, J.C. (1977) Antibodies to major histocompatibility antigens produced by hybrid cell lines. Nature 266, 550–552
Horibata, K., Harris, A.W. (1970) Mouse myelomas and lymphomas in culture. Exp. Cell Res. 60, 61–70
Horisberger, M., Rouvet-Vauthey, M. (1985) Cell wall architecture of the fission yeast Schizosaccharomyces pombe. Experientia 41, 748–750
Horisberger, M., Rouvet-Vauthey, M., Richli, U., Farr, D.A. (1985) Cell wall architecture of the halophilic yeast Saccharomyces rouxii. An immunocytochemical study. Eur. J. Cell Biol. 37, 70–77
Kishida, E., Sone, Y., Shibata, S., Misaki, A. (1989) Preparation and immunochemical characterization of antibody to branched β-(1→3)-d-glucan of Volvariella volvacea, and its use in studies of antitumor actions. Agric. Biol. Chem. 53, 1849–1859
Latge, J.P., Cole, G.T., Horisberger, M., Prevost, M.G. (1986) Ultrastructure and chemical composition of the ballistospore wall of Conidiobolus obscurus. Exp. Mycol. 10, 99–113
Lowry, O.H., Rosenbrough, N.J., Farr, A.L., Randall, R.J. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193, 265–275
Northcote, D.H., Davey, R., Lay, J. (1989) Use of antisera to localize callose, xylan and arabinogalactan in the cell plate, primary and secondary walls of plant cells. Planta 178, 353–366
Oi, V., Herzenberg, L. (1980) In: Selected methods in cellular immunology, pp 351–372, Shigii, S.M., Mishell, B.B. eds. Freeman, San Francisco
Peat, S., Whelan, W.J., Lawley, H.G. (1958) The structure of laminarin. Part II. The minor structural features. J. Chem. Soc. 1, 729–737
Rae, A., Harris, P.J., Bacic, A., Clarke, A.E. (1985) Composition of the cell walls of Nicotiana alata Link et Otto pollen tubes. Planta 166, 128–133
Roy, R., Katzenellenbogen, E., Jennings, H.J. (1984) Improved procedures for the conjugation of oligosaccharides to protein by reductive amination. Can. J. Biochem. Cell Biol. 62, 270–275
Salema, R., Brandao, I. (1973) The use of PIPES buffer in the fixation of plant cells for electron microscopy. J. Submicro. Cytol. 5, 79–96
Spurr, A.R. (1969) A low viscosity resin embedding medium for electron microscopy. J. Ultrastruct. Res. 26, 31–65
Stone, B.A., Evans, N.A., Bonig, I., Clarke, A.E. (1984) The application of Sirofluor, a chemically defined fluorochrome from aniline blue for the histochemical detection of callose. Protoplasma 122, 191–195
Woodward, J.W., Fincher, G.B., Stone, B.A. (1983) Water soluble (1→3), (1→4)-β-d-glucans from barley (Hordeum vulgare) endosperm II. Fine structure. Carbohydr. Polymers 3, 207–226
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P.J.M. is an Australian Postdoctoral Research Fellow. We wish to thank Joan Hoogenraad for her technical assistance with the tissue culture, and Althea Wright for her assistance in the preparation of this paper.
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Meikle, P.J., Bonig, I., Hoogenraad, N.J. et al. The location of (1→3)-β-glucans in the walls of pollen tubes of Nicotiana alata using a (1→3)-β-glucan-specific monoclonal antibody. Planta 185, 1–8 (1991). https://doi.org/10.1007/BF00194507
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DOI: https://doi.org/10.1007/BF00194507