Escobar A L, Cifuentes F, Vergara J L
Department of Physiology, UCLA School of Medicine 90024, USA.
FEBS Lett. 1995 May 15;364(3):335-8. doi: 10.1016/0014-5793(95)00425-9.
A confocal spot detection optical setup was used to record fluorescence signals in response to calcium pulses, elicited by flash photolysis of DM-nitrophen, with the calcium indicators CaOrange-5N and Fluo-3. Our results yield the following conclusions: [Ca2+] changes are almost perfect spikes at pCa 9 and broader transients followed by a step at pCa 7. The [Ca2+] spikes were used to measure the dissociation rate constant of the Ca2+ dyes. Experiments at pCa 7 were used to verify the kinetic rate constants of the dyes and to obtain those of DM-nitrophen. The association rate constant of this compound was found to be more than one order of magnitude faster than that suggested previously. CaOrange-5N was able to track changes in [Ca2+] more accurately than Fluo-3. This latter dye introduced severe distortions which preclude a quantitative deconvolution of the fluorescence transients into changes in the free [Ca2+].
使用共聚焦光斑检测光学装置记录荧光信号，该信号是由DM-硝基苯酚的闪光光解引发的钙脉冲响应信号，使用钙指示剂CaOrange-5N和Fluo-3。我们的结果得出以下结论：在pCa 9时，[Ca2+]变化几乎是完美的尖峰，在pCa 7时是更宽的瞬变，随后是一个阶跃。[Ca2+]尖峰用于测量Ca2+染料的解离速率常数。在pCa 7下进行的实验用于验证染料的动力学速率常数并获得DM-硝基苯酚的动力学速率常数。发现该化合物的缔合速率常数比先前建议的值快一个多数量级。CaOrange-5N比Fluo-3能够更准确地跟踪[Ca2+]的变化。后一种染料引入了严重的失真，这使得无法将荧光瞬变定量反卷积为游离[Ca2+]的变化。
