期刊
JOURNAL OF IMMUNOLOGICAL METHODS
卷 311, 期 1-2, 页码 220-225出版社
ELSEVIER
DOI: 10.1016/j.jim.2006.02.005
关键词
flow cytometry; calcium channels; calcium flux; Indo-1; Fura Red; fluo-3
Indo-1 and high-power water-cooled lasers have been the standard for flow cytometric based Ca2+ flux measurements. With advances in technology and the availability of low-power air-cooled lasers, there is interest in alternative protocols. Here, we have compared Indo-1 with the combination of fluo-3 and Fura Red calcium indicator dyes using low-power air-cooled lasers as the excitation source. The reagents were examined in parallel to detect Ca2+ flux in peripheral blood T lymphocytes and in a T lymphoblastoid cell line. Ca2+ flux was detected with a FACSVantage SE equipped with an Omnichrome Series 74 Helium-Cadmium, or a Spectra Physics 177-G1202 Argon ion air-cooled laser. Following determination of optimal loading conditions, Ca2+ flux was examined in response to membrane receptor stimulation or intracellular Ca2+ mobilization. Dose dependent Ca2+ flux to anti-CD3 and thapsigargin was detected with either Indo-1 or with fluo-3 and Fura Red. The profile of the Ca2+ flux detected by Indo-1 or with fluo-3 and Fura Red appeared similar, with the combination of fluo-3 and Fura Red more sensitive under the particular test conditions. The results clearly demonstrated that Indo-1 could be usefully excited with a low-power air-cooled laser. The alternative use of fluo-3 and Fura Red does not require the availability of a UV capable laser and produced equivalent data. (c) 2006 Elsevier B.V. All rights reserved.
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