Flow cytometric cytotoxicity assay for measuring mammalian and avian NK cell activity. 2002

Csaba Vizler, and Tünde Nagy, and Erzsébet Kusz, and Hristos Glavinas, and Erno Duda
Institute of Biochemistry, Biological Research Center of the Hungarian Academy of Sciences, POB 521, 6701 Szeged, Hungary. vizler@nucleus.szbk.u-szeged.hu

BACKGROUND Flow-cytometric assays are convenient alternatives to classic radioactive natural killer (NK) tests. MitoTracker Green FM, a green fluorescent intracellular probe serving originally for staining mitochondria, seemed especially suitable for labeling NK target cells. Materials and Methods NK target cells were labeled with MitoTracker Green FM. After incubation with effector spleen cells, cell suspensions were stained with propidium iodide (PI), and flow-cytometric analysis was performed. RESULTS MitoTracker Green FM stained efficiently each cell type we assayed, including resting cells, and it was not released from dead cells. NK assays were set up using mouse spleen effector cells and K562 NK target cells. MitoTracker Green FM and PI double staining allowed a discrimination of live and dead target cells, and the cytotoxicity values were in the expected range. Then the method was applied to a less well-known chicken model. We found that chicken-skin fibroblasts had a definite sensitivity to autologous splenic NK cells, sometimes as high as the sensitivity of classic NK targets. CONCLUSIONS Convenient flow-cytometric NK tests can be performed by MitoTracker Green FM and PI staining. Using this method, we demonstrated that chicken fibroblasts are sensitive to the cytotoxic effect of autologous NK cells.

UI MeSH Term Description Entries
D007694 Killer Cells, Natural Bone marrow-derived lymphocytes that possess cytotoxic properties, classically directed against transformed and virus-infected cells. Unlike T CELLS; and B CELLS; NK CELLS are not antigen specific. The cytotoxicity of natural killer cells is determined by the collective signaling of an array of inhibitory and stimulatory CELL SURFACE RECEPTORS. A subset of T-LYMPHOCYTES referred to as NATURAL KILLER T CELLS shares some of the properties of this cell type. NK Cells,Natural Killer Cells,Cell, NK,Cell, Natural Killer,Cells, NK,Cells, Natural Killer,Killer Cell, Natural,NK Cell,Natural Killer Cell
D008810 Mice, Inbred C57BL One of the first INBRED MOUSE STRAINS to be sequenced. This strain is commonly used as genetic background for transgenic mouse models. Refractory to many tumors, this strain is also preferred model for studying role of genetic variations in development of diseases. Mice, C57BL,Mouse, C57BL,Mouse, Inbred C57BL,C57BL Mice,C57BL Mice, Inbred,C57BL Mouse,C57BL Mouse, Inbred,Inbred C57BL Mice,Inbred C57BL Mouse
D002645 Chickens Common name for the species Gallus gallus, the domestic fowl, in the family Phasianidae, order GALLIFORMES. It is descended from the red jungle fowl of SOUTHEAST ASIA. Gallus gallus,Gallus domesticus,Gallus gallus domesticus,Chicken
D003603 Cytotoxins Substances that are toxic to cells; they may be involved in immunity or may be contained in venoms. These are distinguished from CYTOSTATIC AGENTS in degree of effect. Some of them are used as CYTOTOXIC ANTIBIOTICS. The mechanism of action of many of these are as ALKYLATING AGENTS or MITOSIS MODULATORS. Cytolysins,Cytotoxic Agent,Cytotoxic Agents,Cytotoxin,Agent, Cytotoxic
D005347 Fibroblasts Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules. Fibroblast
D005434 Flow Cytometry Technique using an instrument system for making, processing, and displaying one or more measurements on individual cells obtained from a cell suspension. Cells are usually stained with one or more fluorescent dyes specific to cell components of interest, e.g., DNA, and fluorescence of each cell is measured as it rapidly transverses the excitation beam (laser or mercury arc lamp). Fluorescence provides a quantitative measure of various biochemical and biophysical properties of the cell, as well as a basis for cell sorting. Other measurable optical parameters include light absorption and light scattering, the latter being applicable to the measurement of cell size, shape, density, granularity, and stain uptake. Cytofluorometry, Flow,Cytometry, Flow,Flow Microfluorimetry,Fluorescence-Activated Cell Sorting,Microfluorometry, Flow,Cell Sorting, Fluorescence-Activated,Cell Sortings, Fluorescence-Activated,Cytofluorometries, Flow,Cytometries, Flow,Flow Cytofluorometries,Flow Cytofluorometry,Flow Cytometries,Flow Microfluorometries,Flow Microfluorometry,Fluorescence Activated Cell Sorting,Fluorescence-Activated Cell Sortings,Microfluorimetry, Flow,Microfluorometries, Flow,Sorting, Fluorescence-Activated Cell,Sortings, Fluorescence-Activated Cell
D000447 Aldehydes Organic compounds containing a carbonyl group in the form -CHO. Aldehyde
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D013154 Spleen An encapsulated lymphatic organ through which venous blood filters.
D014407 Tumor Cells, Cultured Cells grown in vitro from neoplastic tissue. If they can be established as a TUMOR CELL LINE, they can be propagated in cell culture indefinitely. Cultured Tumor Cells,Neoplastic Cells, Cultured,Cultured Neoplastic Cells,Cell, Cultured Neoplastic,Cell, Cultured Tumor,Cells, Cultured Neoplastic,Cells, Cultured Tumor,Cultured Neoplastic Cell,Cultured Tumor Cell,Neoplastic Cell, Cultured,Tumor Cell, Cultured

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