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Protoplasma

, Volume 148, Issue 2–3, pp 80–86 | Cite as

Cortical fine actin filaments in higher plant cells visualized by rhodamine-phalloidin after pretreatment with m-maleimidobenzoyl N-hydroxysuccinimide ester

  • Seiji Sonobe
  • Hiroh Shibaoka
Article

Summary

Actin filaments in cultured tobacco cells were stained by rhodamine-phalloidin after pretreatment with 100 μM m-maleidobenzoyl N-hydroxysuccinimide ester (MBS) followed by formaldehyde fixation. The use of MBS prior to formaldehyde fixation enabled us to visualize fine, transversely arranged cortical actin filaments in a majority of interphase tobacco cells. It also enabled us to double-stain fine actin filaments and microtubules in the same cells. The pattern of actin filaments and that of microtubules in the cortical region of a single tobacco cell bore a close resemblance to each other. The method which employed MBS was found to be useful also in visualizing fine cortical actin filaments in inner epidermal cells of onion bulbs.

Rhodamine-phalloidin seemed to induce the bundling of actin filaments both tobacco cells and in onion cells when it was applied to the cells which had not been subjected to fixation, indicating that the application of fluorescent-dye-labeled phallotoxins to unfixed cells involves the risk of observing artifically bundled actin filaments.

Keywords

Protein cross-linking reagent Actin filaments Fluorescence microscopy m-Maleimidobenzoyl N-hydroxysuccinimide ester Protein fixation Tobacco BY-2 cell 

Abbreviations

EGTA

ethylenglycol-bis (β-aminoethyl ether)N,N,N′,N′-tetraacetic acid

DMSO

dimethyl sulfoxide

MBS

m-maleimidobenzoyl N-hydroxysuccinimide ester

PBS

phosphate buffer saline

RP

rhodamine-labeled phalloidin

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Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • Seiji Sonobe
    • 1
  • Hiroh Shibaoka
    • 1
  1. 1.Department of Biology, Faculty of ScienceOsaka UniversityJapan

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