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Development of a novel flow cytometric cell-mediated cytotoxicity assay using the fluorophores PKH-26 and TO-PRO-3 iodide

Record type: Article

A flow cytometric (FCM) assay has been developed for the determination of cell-mediated cytotoxicity (CMC). In the assay, the target tumour cell population was labelled with a membrane dye, PKH-26, prior to incubation with splenocyte effector cells. Cell death within the target population was assessed by the addition of the viability probe TO-PRO-3 iodide (TP3) and analysed by flow cytometry. The extent of cytotoxicity was determined by the relative number of live target cells labelled with PKH-26 only and dead, permeabilised cells labelled with both PKH-26 and TP3. This CMC method allows the analysis to be conducted on a single cell basis and overcomes the need for radiochemicals. This communication indicates that the FCM assay is an accurate and reproducible experimental system capable of analysing natural killer (NK) cell and antibody-dependent cell-mediated cytotoxicity. The procedure is comparable to the chromium release assay. We believe that this is one of the first demonstrations of an FCM-based antibody-dependent cell-mediated cytotoxicity (ADCC) assay.

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Citation

Lee-MacAry, Alice E., Ross, Elizabeth L., Davies, Derek, Laylor, Ruthline, Honeychurch, Jamie, Glennie, Martin J., Snary, David and Wilkinson, Robert W. (2001) Development of a novel flow cytometric cell-mediated cytotoxicity assay using the fluorophores PKH-26 and TO-PRO-3 iodide Journal of Immunological Methods, 252, (1-2), pp. 83-92. (doi:10.1016/S0022-1759(01)00336-2).

More information

Published date: 2001
Keywords: cell-mediated cytotoxicity, flow cytometry, PKH-26, TO-PRO-3 iodide

Identifiers

Local EPrints ID: 26434
URI: http://eprints.soton.ac.uk/id/eprint/26434
ISSN: 0022-1759
PURE UUID: b0035ad0-7e59-4657-aa64-ceb11dab8b90

Catalogue record

Date deposited: 24 Apr 2006
Last modified: 17 Jul 2017 16:07

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Contributors

Author: Alice E. Lee-MacAry
Author: Elizabeth L. Ross
Author: Derek Davies
Author: Ruthline Laylor
Author: Jamie Honeychurch
Author: David Snary
Author: Robert W. Wilkinson

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