By David G. Lambert
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Extra resources for Calcium Signaling Protocols - 2nd ed (Methods in Molecular Biology)
1C). Fura-2 Measurements in Cell Suspensions 41 Fig. 1. Carbachol increases [Ca2+]i in suspensions of SH-SY5Y cells. (A) Emission at 340 and 380 nm excitation. Note the antiparallel movement of both traces. (B) Derived 340/380 ratio and (C) [Ca2+]i after calibration. 39, respectively. 18 arbitrary units Շ2% of cell signal. 2. K+ Stimulation in NG108-15 Cells The authors have previously reported a nifedipine sensitive increase in [Ca2+]i in NG108-15 cells in response to depolarization with high K+ (6).
And Pozzan, T. (1987) Fura-2 measurements of cytosolic free Ca2+ in monolayers and suspensions of various types of animal cells. J. Cell Biol. 105, 2719–2727. 27. Merritt, J. , and Hallam, T. J. (1988) Use of manganese to discriminate between calcium influx and mobilization from internal stores in stimulated human neutrophils. J. Biol. Chem. 264, 1522–1527. 28. Rink, T. J. and Cobbold, P. H. (1987) Fluorescence and bioluminescence measurement of cytoplasmic free calcium. Biochem. J. 248, 313–328.
In comparison, recombinant protein probes require a suitable gene expression system. Plasmid transfection techniques are not readily suited for freshly isolated or quiesencent cells. On the other hand viral vectors offer much wider access of the probes to cells and tissues. 1, there is also delay in the order of 24 h between infection and measurement. For freshly isolated cells this delay may be reduced if the probe can be introduced into the tissue sometime before the cells are isolated. As ever, choosing a probe depends on the specific aims and objectives of the experiment.
Calcium Signaling Protocols - 2nd ed (Methods in Molecular Biology) by David G. Lambert