Emission ratiometry for simultaneous calcium and action potential measurements with coloaded dyes in rabbit hearts: Reduction of motion and drift
Emission ratiometry for simultaneous calcium and action potential measurements with coloaded dyes in rabbit hearts: Reduction of motion and drift
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DOI：
10.1046/j.1540-8167.2003.02077.x
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发表时间：
2003-01-01
影响因子：
2.7
通讯作者：
Knisley, SB
中科院分区：
文献类型：
--
作者：
Kong, W;Walcott, GP;Knisley, SB
介绍：心脏钙瞬态（Ca）和跨膜动作电位（AP）的光学测量可以与发射比测定同时进行，以减少运动伪影和光漂白效应。我们检测了灌注了Rh237和绿色Ca染料（Fluo-4或Oregon Green BAPTA 1）的兔心脏的发射光谱变化，以确定发射比率测定法的波长范围，并测试比率测定法是否能减少运动伪影和漂移。方法和结果：用488-mn激光照射心脏，用荧光分光计收集489 - 838 nm 1 kHz频率的荧光。以663 ~ 685 nm （IS）为分母，以510 ~ 532 nm为分子的Ca敏感波段为分子的比率测定法减少了运动伪像，量化为Ca的相对振幅增加了1.4倍（P < 0.05）。以AP敏感波段772 ~ 794 nm为分母，以IS为分子进行比率测定，AP相对振幅增加1.7倍（P < 0.05）。对Ca和AP的比值测定可减少光漂依赖性漂移，分别为0.6 （P < 0.05）和0.45 （P < 0.05）。结论：同时测定Ca和AP的比值可减少染色心脏的运动伪影和漂移。
Introduction: Optical measurements of the cardiac calcium transient (Ca) and transmembrane action potential (AP) may be performed simultaneously with emission ratiometry to lessen motion artifacts and photobleaching effects. We examined changes in emission spectrum in perfused rabbit hearts coloaded with Rh237 and a green-emitting Ca dye (Fluo-4 or Oregon Green BAPTA 1) to determine wavelength bands for emission ratiometry and to test whether ratiometry reduces motion artifacts and drift.Methods and Results: A 488-mn laser illuminated hearts while a spectrofluorometer collected fluorescence from 489 to 838 nm at 1 kHz. Ratiometry with the Ca- and AP-insensitive emission band 663 to 685 nm (IS) as denominator and the Ca-sensitive band 510 to 532 nm as numerator lessened motion artifacts, which was quantified as a 1.4-fold increase in relative amplitude of the Ca (P < 0.05). Ratiometry with the AP-sensitive band 772 to 794 nm as denominator and the IS as numerator produced a 1.7-fold increase in relative amplitude of the AP (P < 0.05). The ratiometry decreased photobleaching-dependent drift by a factor of 0.6 (P < 0.05) for Ca and 0.45 (P < 0.05) for AP.Conclusion: Simultaneous Ca and AP emission ratiometry reduces motion artifacts and drift in hearts with coloaded dyes.