Single-colour flow cytometric assay to determine NK cell-mediated cytotoxicity and viability against non-adherent human tumor cells |
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| Authors: | Ajit Thakur Abeyat Zaman Jeff Hummel Kim Jones Gonzalo Hortelano |
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| Affiliation: | (1) School of Biomedical Engineering, McMaster University, 5380 Glen Erin Dr., Mississauga, ON, L5M5C7, Canada;(2) Bachelor of Health Sciences Program, McMaster University, 1200 Main Street, Hamilton, ON, L8N 3Z5, Canada;(3) Centre for Gene Therapeutics, McMaster University, MDCL Building, Rm. 5070, 1200 Main Street, Hamilton, ON, L8N 3Z5, Canada;(4) Department of Chemical Engineering, McMaster University, Hamilton, ON, L8N 3Z5, Canada;(5) Department of Pathology & Molecular Medicine, McMaster University, Hamilton, ON, L8N 3Z5, Canada |
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| 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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