کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
1177063 961937 2009 9 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
A nonfluorescent, broad-range quencher dye for Förster resonance energy transfer assays
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
A nonfluorescent, broad-range quencher dye for Förster resonance energy transfer assays
چکیده انگلیسی

We report here a novel, water-soluble, nonfluorescent dye that efficiently quenches fluorescence from a broad range of visible and near-infrared (NIR) fluorophores in Förster resonance energy transfer (FRET) systems. A model FRET-based caspase-3 assay system was used to test the performance of the quencher dye. Fluorogenic caspase-3 substrates were prepared by conjugating the quencher, IRDye® QC-1, to a GDEVDGAK peptide in combination with fluorescein (emission maximum ∼540 nm), Cy3 (∼570 nm), Cy5 (∼670 nm), IRDye 680 (∼700 nm), IRDye 700DX (∼690 nm), or IRDye 800CW (∼790 nm). The Förster distance R0 values are calculated as 41 to 65 Å for these dye/quencher pairs. The fluorescence quenching efficiencies of these peptides were determined by measuring the fluorescence change on complete cleavage by recombinant caspase-3 and ranged from 97.5% to 98.8%. The fold increase in fluorescence on caspase cleavage of the fluorogenic substrates ranged from 40 to 83 depending on the dye/quencher pair. Because IRDye QC-1 effectively quenches both the NIR fluorophores (e.g., IRDye 700DX, IRDye 680, IRDye 800CW) and the visible fluorophores (e.g., fluorescein, Cy3, Cy5), it should find broad applicability in FRET assays using a wide variety of fluorescent dyes.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytical Biochemistry - Volume 388, Issue 2, 15 May 2009, Pages 220–228
نویسندگان
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