Flow cytometric measurement of [Ca2+]i and pHi in conjugated natural killer cells and K562 target cells during the cytotoxic process

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Graft, Marja van and Kraan, Yvonne M. and Segers-Nolten, Ine M.J. and Radosevic, Katarina and Grooth, Bart G. de and Greve, Jan (1993) Flow cytometric measurement of [Ca2+]i and pHi in conjugated natural killer cells and K562 target cells during the cytotoxic process. Cytometry Part A, 14 (3). pp. 257-264. ISSN 0196-4763

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Abstract:We describe a flow cytometric assay that enables one to follow conjugate formation between cytotoxic cells and their target cells during the cytotoxic process. In addition, the internal calcium concentration ([Ca2+]i) and internal pH (pHi) of the conjugated cells can be monitored and directly compared to the nonconjugated cells. This is achieved by labeling one cell type with the Ca2+-specific dye Fluo-3, while the other cell type is labeled with the pH-sensitive dye SNARF-1. As these fluorochromes have different emission spectra, events positive for both fluorochromes are identified as conjugates. The results show that the conjugates can be clearly distinguished from single cytotoxic cells [natural killer (NK) cells] and target cells [K562 cells, (TC)]. Upon binding, [Ca2+]i is increased in the NK cells as well as in the TC. In conjugated NK cells this increase of [Ca2+]i is temperature dependent and is followed by a decrease to a normal [Ca2+]i value later on. The [Ca2+]i in NK cells increases in 2 steps, which may be related to the binding-and lethal hit phase. Upon conjugate formation, NK cells show a slight increase in pHi (0.2-0.3 pH units). TC do not reveal a significant change in pHi.
Item Type:Article
Copyright:© 1993 Wiley-Liss
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Science and Technology (TNW)
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Link to this item:http://purl.utwente.nl/publications/60718
Official URL:http://dx.doi.org/10.1002/cyto.990140304
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