Characterization of Acanthamoeba-microsphere association by multiparameter flow cytometry and confocal microscopy

Edward A G Elloway, Roger A. Bird, Christopher J. Hewitt, Steven L. Kelly, Stephen N. Smith*

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

Background: Acanthamoebae, in common with other protozoa, readily endocytose particulate material, which in turn may lead to the spread of infectious disease. Methods: Evaluation and quantification of plain and carboxylate FITC-microsphere association with acanthamoebal trophzoites was undertaken using a combination of flow cytometry and confocal microscopy. Trophozoites from strains and species of Acanthamoeba were exposed to plain and carboxylate FITC-microspheres. Microsphere size and aspects such as trophozoite starvation, maturity, and exposure to metabolic inhibitors were assessed. Results: All species and strains of Acanthamoeba readily endocytosed plain and carboxylate microspheres. Starving trophozoites significantly increased binding and potential ingestion of microspheres, whereas trophozoites of increasing maturity lost such abilities. Trophozoites showed a significant preference for 2.0- and 3.0-μm-diameter microspheres when compared with other sizes, which in turn could occupy much of the cytoplasm. The physiological inhibitors sodium azide, 2,4-clinitrophenol, and cytochalasin B reduced microsphere association with trophozoites; however, some microspheres still bound and associated with trophozoites after inhibitor exposure, a manifestation of both active and inactive agent involvement in microsphere endocytosis. Conclusions: Even though the origins of microsphere binding by acanthamoebal trophozoite remains shrouded, the combination of flow cytometry and confocal microscopy supported synergistic quantification and qualification of trophozoite-microsphere endocytosis.

Original languageEnglish
Pages (from-to)266-272
Number of pages7
JournalCytometry: Part A
Volume69A
Issue number4
Early online date23 Feb 2006
DOIs
Publication statusPublished - Apr 2006

Fingerprint

Acanthamoeba
Trophozoites
Microspheres
Confocal Microscopy
Flow Cytometry
Endocytosis
Fluorescein-5-isothiocyanate
Sodium Azide
Cytochalasin B
Starvation
Communicable Diseases
Cytoplasm
Eating

Keywords

  • acanthamoeba
  • endocytosis
  • FITC-microsphere
  • trophozoite

Cite this

Elloway, Edward A G ; Bird, Roger A. ; Hewitt, Christopher J. ; Kelly, Steven L. ; Smith, Stephen N. / Characterization of Acanthamoeba-microsphere association by multiparameter flow cytometry and confocal microscopy. In: Cytometry: Part A. 2006 ; Vol. 69A, No. 4. pp. 266-272.
@article{ab4a906fac9b47f3aa960d1aaf040b5b,
title = "Characterization of Acanthamoeba-microsphere association by multiparameter flow cytometry and confocal microscopy",
abstract = "Background: Acanthamoebae, in common with other protozoa, readily endocytose particulate material, which in turn may lead to the spread of infectious disease. Methods: Evaluation and quantification of plain and carboxylate FITC-microsphere association with acanthamoebal trophzoites was undertaken using a combination of flow cytometry and confocal microscopy. Trophozoites from strains and species of Acanthamoeba were exposed to plain and carboxylate FITC-microspheres. Microsphere size and aspects such as trophozoite starvation, maturity, and exposure to metabolic inhibitors were assessed. Results: All species and strains of Acanthamoeba readily endocytosed plain and carboxylate microspheres. Starving trophozoites significantly increased binding and potential ingestion of microspheres, whereas trophozoites of increasing maturity lost such abilities. Trophozoites showed a significant preference for 2.0- and 3.0-μm-diameter microspheres when compared with other sizes, which in turn could occupy much of the cytoplasm. The physiological inhibitors sodium azide, 2,4-clinitrophenol, and cytochalasin B reduced microsphere association with trophozoites; however, some microspheres still bound and associated with trophozoites after inhibitor exposure, a manifestation of both active and inactive agent involvement in microsphere endocytosis. Conclusions: Even though the origins of microsphere binding by acanthamoebal trophozoite remains shrouded, the combination of flow cytometry and confocal microscopy supported synergistic quantification and qualification of trophozoite-microsphere endocytosis.",
keywords = "acanthamoeba, endocytosis, FITC-microsphere, trophozoite",
author = "Elloway, {Edward A G} and Bird, {Roger A.} and Hewitt, {Christopher J.} and Kelly, {Steven L.} and Smith, {Stephen N.}",
year = "2006",
month = "4",
doi = "10.1002/cyto.a.20210",
language = "English",
volume = "69A",
pages = "266--272",
journal = "Cytometry: Part A",
issn = "1552-4922",
publisher = "Wiley-Liss Inc.",
number = "4",

}

Characterization of Acanthamoeba-microsphere association by multiparameter flow cytometry and confocal microscopy. / Elloway, Edward A G; Bird, Roger A.; Hewitt, Christopher J.; Kelly, Steven L.; Smith, Stephen N.

In: Cytometry: Part A, Vol. 69A, No. 4, 04.2006, p. 266-272.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Characterization of Acanthamoeba-microsphere association by multiparameter flow cytometry and confocal microscopy

AU - Elloway, Edward A G

AU - Bird, Roger A.

AU - Hewitt, Christopher J.

AU - Kelly, Steven L.

AU - Smith, Stephen N.

PY - 2006/4

Y1 - 2006/4

N2 - Background: Acanthamoebae, in common with other protozoa, readily endocytose particulate material, which in turn may lead to the spread of infectious disease. Methods: Evaluation and quantification of plain and carboxylate FITC-microsphere association with acanthamoebal trophzoites was undertaken using a combination of flow cytometry and confocal microscopy. Trophozoites from strains and species of Acanthamoeba were exposed to plain and carboxylate FITC-microspheres. Microsphere size and aspects such as trophozoite starvation, maturity, and exposure to metabolic inhibitors were assessed. Results: All species and strains of Acanthamoeba readily endocytosed plain and carboxylate microspheres. Starving trophozoites significantly increased binding and potential ingestion of microspheres, whereas trophozoites of increasing maturity lost such abilities. Trophozoites showed a significant preference for 2.0- and 3.0-μm-diameter microspheres when compared with other sizes, which in turn could occupy much of the cytoplasm. The physiological inhibitors sodium azide, 2,4-clinitrophenol, and cytochalasin B reduced microsphere association with trophozoites; however, some microspheres still bound and associated with trophozoites after inhibitor exposure, a manifestation of both active and inactive agent involvement in microsphere endocytosis. Conclusions: Even though the origins of microsphere binding by acanthamoebal trophozoite remains shrouded, the combination of flow cytometry and confocal microscopy supported synergistic quantification and qualification of trophozoite-microsphere endocytosis.

AB - Background: Acanthamoebae, in common with other protozoa, readily endocytose particulate material, which in turn may lead to the spread of infectious disease. Methods: Evaluation and quantification of plain and carboxylate FITC-microsphere association with acanthamoebal trophzoites was undertaken using a combination of flow cytometry and confocal microscopy. Trophozoites from strains and species of Acanthamoeba were exposed to plain and carboxylate FITC-microspheres. Microsphere size and aspects such as trophozoite starvation, maturity, and exposure to metabolic inhibitors were assessed. Results: All species and strains of Acanthamoeba readily endocytosed plain and carboxylate microspheres. Starving trophozoites significantly increased binding and potential ingestion of microspheres, whereas trophozoites of increasing maturity lost such abilities. Trophozoites showed a significant preference for 2.0- and 3.0-μm-diameter microspheres when compared with other sizes, which in turn could occupy much of the cytoplasm. The physiological inhibitors sodium azide, 2,4-clinitrophenol, and cytochalasin B reduced microsphere association with trophozoites; however, some microspheres still bound and associated with trophozoites after inhibitor exposure, a manifestation of both active and inactive agent involvement in microsphere endocytosis. Conclusions: Even though the origins of microsphere binding by acanthamoebal trophozoite remains shrouded, the combination of flow cytometry and confocal microscopy supported synergistic quantification and qualification of trophozoite-microsphere endocytosis.

KW - acanthamoeba

KW - endocytosis

KW - FITC-microsphere

KW - trophozoite

UR - http://www.scopus.com/inward/record.url?scp=33645946698&partnerID=8YFLogxK

UR - http://onlinelibrary.wiley.com/doi/10.1002/cyto.a.20210/abstract

U2 - 10.1002/cyto.a.20210

DO - 10.1002/cyto.a.20210

M3 - Article

C2 - 16498687

AN - SCOPUS:33645946698

VL - 69A

SP - 266

EP - 272

JO - Cytometry: Part A

JF - Cytometry: Part A

SN - 1552-4922

IS - 4

ER -