کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
9743780 | 1491199 | 2005 | 10 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Interferences in Fluo-3 based ion-flux assays for ligand-gated-ion channels
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
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چکیده انگلیسی
1-[2-Amino-5-(2,7-dicholoro-6-hydoxy-3-oxo-3H-xanthen-9-yl)phenoxy]-2-(2â²-amino-5â²methylphenoxylethane)-N,N,Nâ²,Nâ²-tetraacetic acid (Fluo-3), a highly sensitive calcium indicator dye, is commonly used for measuring changes in intracellular Ca2+ concentrations in response to cellular ion-channel function. In particular, Fluo-3 has been used extensively for in vivo monitoring of the response of ligand-gated-ion-channels (LGIC), such as the nicotinic acetylcholine receptor (nAChR), to the presence of agonists and antagonists. Our initial goal was to use Fluo-3 for in vitro measurement of Ca2+ influx across artificial liposomes containing reconstituted nAChR. However, it became apparent during initial control experiments that agonist and antagonist-like responses could be obtained for several known nAChR effectors in the presence of the dye alone. Steady-state and time-resolved fluorescence measurements of Fluo-3 in the presence of various nAChR agonists and antagonists revealed that Fluo-3 formed a ground state complex with the antagonist d-tubocurarine, leading to a decrease in emission intensity similar to that expected upon blockage of the ion-channel. More surprisingly, it was found that nicotine, an agonist of nAChR, could disrupt the formation of this complex with a concomitant reversal of the static quenching effects, leading to increased emission which was consistent with the expected changes in intensity for a channel opening event. These results demonstrate that interactions of agonists and antagonists with Fluo-3 could lead to false positives in screens of ligand-gated-ion-channels such as nAChR.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytica Chimica Acta - Volume 537, Issues 1â2, 29 April 2005, Pages 125-134
Journal: Analytica Chimica Acta - Volume 537, Issues 1â2, 29 April 2005, Pages 125-134
نویسندگان
Travis R. Besanger, Hitesh Bhanabhai, John D. Brennan,