Advertisement

Planta

, Volume 168, Issue 4, pp 471–481 | Cite as

Immunoelectron-microscopy localization of abscisic acid with colloidal gold on Lowicryl-embedded tissues of Chnopodium polyspermum L.

  • L. Sossountzov
  • B. Sotta
  • R. Maldiney
  • I. Sabbagh
  • E. Miginiac
Article

Abstract

Further study on the localization of abscisic acid (ABA) has been undertaken at the ultrastructural level in Chenopodium polyspermum L. Axillary-bud-bearing nodes on the main axis were fixed with soluble 1-(3-dimethylaminopropyl)-3 ethyl carbodiimide, then postfixed with paraformaldehyde and embedded in Lowicryl K4M at-20° C. Ultrathin sections mounted on grids were successively incubated with rabbit anti-ABA antibodies and with gold-labelled goat anti-rabbit anti-bodies (40 nm particle size). Control sections treated with preimmune rabbit serum and ABA-preabsorbed antibodies were devoid of label. The background staining was very low with this technique. Quantitative analysis of the immunolabelling showed that two main sites of ABA accumulation could be defined: first, plastids in cortical cells and vascular parenchyma cells associated with sieve elements and xylem vessels; second, the cell cytoplasm and nucleus in the axillary bud tip and in procambial strands. In vascular bundles, the cambial cells showed no immunoreactivity. These observations support the hypothesis for the cytoplasmic synthesis of ABA which is subsequently trapped in plastids as cells mature.

Key words

Abscisic acid (localization) Chenopodium Immunogold electron microscopy Lowicryl K4M 

Abbreviations

ABA

abscisic acid

EDC

1-(3-dimethyl-aminopropyl)-3-ethyl carbodiimide

GAR 40

goat anti-rabbit antibodies labelled with colloidal gold of particle size 40 nm

IgG

immunoglobulin G

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Bendayan, M. (1984) Protein A-gold electron microscopic immunocytochemistry: methods, applications and limitations. J. Electron Microsc. Technol. 1, 243–270Google Scholar
  2. Carlemalm, E., Garavito, R.M., Villiger, W. (1982) Resin development for electron microscopy and an analysis of embedding at low temperature. J. Microsc. (Oxford) 126, 123–143Google Scholar
  3. Cowan, I.R., Raven, J.A., Hartung, W., Farquhar, G.D. (1982) A possible role for abscisic acid in coupling stomatal conductance and photosynthetic carbon metabolism in leaves. Aust. J. Plant Physiol. 9, 489–498Google Scholar
  4. Craig, S., Goodchild, D.J. (1982) Post-embedding immunolabelling. Some effects of tissue preparation on the antigenicity of plant proteins. Eur. J. Cell Biol. 28, 251–256Google Scholar
  5. Craig, S., Miller, C. (1984) LR white resin and improved ongrid immunogold detection of Vicilin, a pea seed storage protein. Cell Biol. Int. Rep. 8, 879–886Google Scholar
  6. De Mey, J. (1983) Colloïdal gold probes in immunocytochemistry. In: Immunocytochemistry: practical applications in pathology and biology, vol 1, pp. 82–113, Polak, J.M., Van Noorden, S., eds. Wright, BristolGoogle Scholar
  7. Goodchild, D.J., Andersson, B., Anderson, J.M. (1985a) Immunocytochemical localization of polypeptides associated with the oxygen evolving system of photosynthesis. Eur. J. Cell. Biol. 36, 294–298Google Scholar
  8. Goodchild, D.J., Anderson, J.M., Andersson, B. (1985b) Immunocytochemical localization of the cytochrome b/f complex of chloroplast thylakoid membranes Cell Biol. Int. Rep. 9, 715–721Google Scholar
  9. Greenwood, J.S., Chrispeels, M.J. (1985) Immunocytochemical localization of phaseolin and phytohemagglutinin in the endoplasmic reticulum and Golgi complex of developing bean cotyledons. Planta 164, 295–302Google Scholar
  10. Harris, N., Croy, R.R.D. (1985) The major albumin protein from pea (Pisum sativum L.). Localization by immunocytochemistry. Planta 165, 522–526Google Scholar
  11. Hartung, W., Heilmann, B., Gimmler, H. (1981) Do chloroplasts play a role in abscisic acid synthesis? Plant Sci. Lett. 22, 235–242Google Scholar
  12. Herman, E.M., Shannon L.M. (1984) Immunocytochemical evidence for the involvement of Golgi apparatus in the deposition of seed lectin of Bauhinia purpura (Leguminosae). Protoplasma 121, 163–170Google Scholar
  13. Leroux, B., Maldiney, R., Miginiac, E., Sossountzov, L., Sotta, B. (1985) Comparative quantitation of abscisic acid in plant extracts by gas-liquid chromotography and an enzyme-linked immunosorbent assay using the avidin-biotin system. Planta 166, 524–529Google Scholar
  14. Maldiney, R., Leroux, B., Sabbagh, I., Sotta, B., Sossountzov, L., Miginiac, E. (1986) Enzyme immunoassay to quantify three phytohormones: auxin, abscisic acid and zeatin-riboside, using a biotin/avidin system. J. Immunol. Meth., in pressGoogle Scholar
  15. Milborrow, B.V. (1979) Antitraspirants and regulation of abscisic acid content. Aust. J. Plant Physiol. 6, 249–254Google Scholar
  16. Newman, G.R., Jasani, B., Williams, E.D. (1983) A simple post-embedding system for the rapid demonstration of tissue antigens under the electron microscope. Histochem. J. 15, 543–555Google Scholar
  17. Rodriguez, D., Nicolas, G., Aldasoro, J.J., Hernandez-Nistal, J., Babiano, M.J., Matilla, A. (1985) Altered development of polysomal RNA activity in chick-pea (Cicer arietimum I.) embryonic axes. Effects of abscisic acid and temperature. Planta 164, 517–523Google Scholar
  18. Roth, J., Bendayan, M., Orci, L. (1978) Ultrastructural localization of intracellular antigens by the use of protein-A gold complex. J. Histochem. Cytochem. 26, 1074–1081Google Scholar
  19. Roth, J., Bendayan, M., Carlemalm, E., Villiger, W., Garavito, M. (1981) Enhancement of structural preservation and immunocytochemical staining in low temperature embedded pancreatic tissue. J. Histochem. Cytochem. 29, 663–671Google Scholar
  20. Shaw, P.J., Henwood, J.A. (1985) Immuno-gold localization of cytochrome f, light-harvesting complex, ATP synthase and ribulose 1,5-bisophosphate carboxylase/oxygenase. Planta 165, 333–339Google Scholar
  21. Sotta, B., Sossountzov, L., Maldiney, R., Sabbagh, I., Tachon, P., Miginiac, E. (1985) Abscisic acid localization by light microscopic immunohistochemistry in Chenopodium polyspermum L. J. Histochem. Cytochem. 33, 201–208Google Scholar
  22. Tomenius, K., Clapham, D., Oxelfelt P. (1983) Localization by immunogold cytochemistry of viral antigens in sections of plant cells infected with red clover mottle virus. J. Gen. Virol. 645, 2669–2678Google Scholar
  23. Walton, D.C. (1980) Biochemistry and physiology of abscisic acid. Annu. Rev. Plant. Physiol. 31, 453–489Google Scholar
  24. Yamamoto, N., Yasuda, K. (1977) Use of a water soluble carbodiimide as a fixing agent. Acta Histochem. Cytochem. 10, 14–37Google Scholar
  25. Zavala, M.E., Brandon, B.L. (1983) Localization of a phytohormone using immunocytochemistry. J. Cell Biol. 97, 1235–1239Google Scholar
  26. Zeevaart, J.A.D. (1977) Sites of abscisic acid synthesis and metabolism in Ricinus communis. L. Plant Physiol. 59, 788–791Google Scholar

Copyright information

© Springer-Verlag 1986

Authors and Affiliations

  • L. Sossountzov
    • 1
  • B. Sotta
    • 1
  • R. Maldiney
    • 1
  • I. Sabbagh
    • 2
  • E. Miginiac
    • 1
  1. 1.Institut de Physiologie Végétale, CNRS, Laboratoire de Physiologie végétale, T 53, 5ème étageUniversité P et M CurieParis Cedex 05France
  2. 2.Domaine du PoirierInstitut Pasteurl'ArbreslesFrance

Personalised recommendations