Abstract
A flow cytometry-based cytotoxicity (FCC) assay was developed using a single fluorophore, calcein-acetoxymethyl diacetylester (calcein-AM), to measure NK cell-mediated cytotoxicity. Non-adherent human K562 and U937 target cells were individually labelled with calcein-AM and co-incubated with effector NK cells to measure calcein loss, and therefore calculate target cell cytotoxicity. This FCC assay also provided a measure of sample viability. Notably, cell viability measured by traditional calcein/7-amino-actinomycin D (7-AAD) double labelling and Trypan Blue methods were comparable to the viability calculated using calcein-loss FCC. This FCC assay may also be used with various effector and target cell types and as a multi-parameter tool to measure viability and immunophenotype cells for tissue engineering purposes.
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Acknowledgments
We would like to thank Dr. Heather Sheardown, Dr. Karen Mossman, Dr. Ali Ashkar, Dr. Firoz Mian, Dr. Iran Rashedi, Glenn McClung, Ruchira Sengupta, Faheem Dinath and Dave Morrison for providing us with materials and technical support, and to the Natural Science and Engineering Research Council (NSERC) for their funding to AT and KJ.
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Thakur, A., Zaman, A., Hummel, J. et al. Single-colour flow cytometric assay to determine NK cell-mediated cytotoxicity and viability against non-adherent human tumor cells. Biotechnol Lett 34, 447–453 (2012). https://doi.org/10.1007/s10529-011-0828-9
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DOI: https://doi.org/10.1007/s10529-011-0828-9