| ImageStream cytometry extends the analysis of phagocytosis and oxidative burst. | |
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MedLine Citation:
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PMID: 21473709 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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Abstract Aim. Phagocytosis is often measured using conventional microscopy and flow cytometry. ImageStream cytometry is a new technology that combines the advantages of both methods, enabling statistically robust microscopic applications. We compared ImageStream cytometry to flow cytometry in a whole blood model of phagocytosis with viable, fluorescence-marked Staphylococcus aureus. We furthermore measured the co-localization of intracellular bacteria to sites of oxidative burst, as well as changes in cell size and actin levels as a result of phagocytosis. Experimental design. Fluorescence-labeled S. aureus in a ratio of 5:1 bacteria per leukocyte were added to whole blood. Phagocytosis was stopped at different time points. After staining of neutrophils and lysis of erythrocytes, samples were analysed by ImageStream cytometry and flow cytometry. Results. Phagocytosis and oxidative burst determined by flow cytometry and ImageStream cytometry showed strong correlation. In contrast to flow cytometry, ImageStream cytometry easily detected and excluded extracellular adherent bacteria from the measurement of phagocytosis, and enumerated the bacteria within each neutrophil. Using the Bright Detail Similarity score, we identified a subset of neutrophils with intracellular bacteria co-localized to sites of oxidative burst activity. Phagocytosis resulted in an increase in cell size and actin polymerization as determined by an increase in phalloidin fluorescence intensity. Conclusions. We describe a simple whole blood image-based method for measuring bacterial phagocytosis and oxidative burst. ImageStream cytometry provides the spatial resolution to determine the number of bacteria ingested and the sub-cellular localization and trafficking patterns that enables a more complete evaluation of the phagocytic process. |
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Authors:
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Annette Ploppa; Thaddeus C George; Klaus E Unertl; Boris Nohe; Marcel E Durieux |
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Publication Detail:
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Type: JOURNAL ARTICLE Date: 2011-4-8 |
Journal Detail:
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Title: Scandinavian journal of clinical and laboratory investigation Volume: - ISSN: 1502-7686 ISO Abbreviation: - Publication Date: 2011 Apr |
Date Detail:
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Created Date: 2011-4-8 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 0404375 Medline TA: Scand J Clin Lab Invest Country: - |
Other Details:
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Languages: ENG Pagination: - Citation Subset: - |
Affiliation:
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Department of Anaesthesiology and Intensive Care Medicine, Eberhard-Karls University, Tuebingen, Germany. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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