A novel approach to dual excitation ratiometric optical mapping of cardiac action potentials with di-4-ANEPPS using pulsed LED excitation.
A novel approach to dual excitation ratiometric optical mapping of cardiac action potentials with di-4-ANEPPS using pulsed LED excitation.
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DOI：
10.1109/tbme.2011.2148719
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发表时间：
2011-07
期刊：
影响因子：
--
通讯作者：
Rogers JM
中科院分区：
文献类型：
--
作者：
Bachtel AD;Gray RA;Stohlman JM;Bourgeois EB;Pollard AE;Rogers JM
We developed a new method for ratiometric optical mapping of transmembrane potential (Vm) in cardiac preparations stained with di-4-ANEPPS. Vm-dependent shifts of excitation and emission spectra establish two excitation bands (<481 and >481 nm) that produce fluorescence changes of opposite polarity within a single emission band (575–620 nm). The ratio of these positive and negative fluorescence signals (excitation ratiometry) increases Vm sensitivity and removes artifacts common to both signals. We pulsed blue (450±10 nm) and cyan (505±15 nm) light emitting diodes (LEDs) at 375 Hz in alternating phase synchronized to a camera (750 frames-per-second). Fluorescence was bandpass filtered (585±20 nm). This produced signals with upright (blue) and inverted (cyan) action potentials (APs) interleaved in sequential frames. In 4 whole swine hearts with motion chemically arrested, fractional fluorescence for blue, cyan, and ratio signals was 1.2±0.3%, 1.2±0.3%, and 2.4±0.6%, respectively. Signal-to-noise ratios were 4.3±1.4, 4.0±1.2, and 5.8±1.9, respectively. After washing out the electromechanical uncoupling agent, we characterized motion artifact by cross-correlating blue, cyan, and ratio signals with a signal with normal AP morphology. Ratiometry improved cross-correlation coefficients from 0.50±0.48 to 0.81±0.25, but did not cancel all motion artifacts. These findings demonstrate the feasibility of pulsed LED excitation ratiometry in myocardium.
