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Please use this identifier to cite or link to this item: http://repository.li.mahidol.ac.th/dspace/handle/123456789/24066
Title: Flow cytometric quantitation of opsonophagocytosis and intracellular killing of Candida albicans using a whole blood microassay
Authors: Kovit Pattanapanyasat
Kasama Sukapirom
Kalaya Tachavanich
Sakorn Kaewmoon
Mahidol University
Keywords: Biochemistry, Genetics and Molecular Biology;Medicine
Issue Date: 1-Dec-2007
Citation: Cytometry Part A. Vol.71, No.12 (2007), 1027-1033
Abstract: Flow cytometric assays for quantifying polymorphonuclear leucocyte (PMN) function involve peripheral blood fractionation methods. However, most are of limited use for conditions like pediatrics and thalassemia, as they may require large blood volumes or complicated by red blood cell contamination. In whole blood assay, 500-μl aliquots of normal or thalassemic whole blood were incubated with bis-carboxyethyl-carboxyfluorescein pentaacetoxymethylester (BCECF-AM)-labeled Candida albicans cells and phycoerythrin-conjugated anti-CD13 monoclonal antibody for PMNs. Phagocytosis was quantitated by gating on CD13 label PMNs and determining the frequency of phagocytosed yeast using flow cytometry. The killing activity was quantitated by estimating the number of viable fluorescence retaining yeast cells liberated following lysis of PMNs. In normal control or thalassemia samples, nonphagocytosed yeast and PMNs with or without phagocytosed yeast were readily distinguished by two-color dot plot, permitting phagocytosis estimates. Viable and nonviable yeast killed by whole blood PMNs were distinguished by side scatter and fluorescence, permitting killing estimates. The patterns seen using whole blood was similar to that seen with fractionated PMNs. Data from thalassemia patients showed similar opsonophagocytosis but slightly decreased intracellular killing of yeast cells when compared with normal subjects. This assay provides an alternative method to assess PMN function in small blood samples, which could be especially useful in conditions like thalassemia and pediatric patients. © 2007 International Society for Analytical Cytology.
URI: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=36849010134&origin=inward
http://repository.li.mahidol.ac.th/dspace/handle/123456789/24066
ISSN: 15524930
15524922
Appears in Collections:Scopus 2006-2010

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