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


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), 83-92. ( doi:10.1016/S0022-1759(01)00336-2).

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Description/Abstract

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.

Item Type: Article
ISSNs: 0022-1759 (print)
Related URLs:
Keywords: cell-mediated cytotoxicity, flow cytometry, PKH-26, TO-PRO-3 iodide
Subjects: R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
Q Science > QR Microbiology > QR180 Immunology
Divisions: University Structure - Pre August 2011 > School of Medicine > Cancer Sciences
ePrint ID: 26434
Date Deposited: 24 Apr 2006
Last Modified: 27 Mar 2014 18:15
Contact Email Address: R.Wilkinson@icrf.icnet.uk
URI: http://eprints.soton.ac.uk/id/eprint/26434

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