Ray Krishanu, Zhang Jian, Lakowicz Joseph R
Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, 725 West Lombard Street, Baltimore, Maryland 21201, USA.
Anal Chem. 2008 Oct 1;80(19):7313-8. doi: 10.1021/ac8009356. Epub 2008 Sep 5.
In this paper, we introduce the use of fluorescence lifetime correlation spectroscopy to study the metal-fluorophore interactions in solution at the single-fluorophore level. A single-stranded oligonucleotide was chemically bound to a 50-nm-diameter single silver particle, and a Cy5-labeled complementary single-stranded oligonucleotide was hybridized with the silver particle-bound oligonucleotide. The distance between the fluorophore and silver particle was maintained by a rigid hybridized DNA duplex of 8 nm in length. The single Cy5-DNA-Ag particles showed more than 10-fold increase in fluorescence intensity and a 5-fold decrease in emission lifetimes as compared with Cy5-DNA free molecules in the absence of metal. The decrease of lifetime for the Cy5-DNA-Ag particle allowed us to resolve the correlation functions of the two species based on the intensity decays. The increased brightness of the Cy5-DNA-Ag particle as compared to free Cy5-DNA resulted in an increased contribution of Cy5-DNA-Ag to the correlation function of the mixture. These results show that the effects of metal particles on fluorophores can be used to detect the small fractional populations of the metal-bound species in the presence of a larger number of less bright species. Our results also suggest that these bright fluorophores conjugated to silver particles could be used as the fluorescent probes for clinical detection in the biological samples with the high background.
在本文中,我们介绍了利用荧光寿命相关光谱法在单荧光团水平上研究溶液中金属 - 荧光团相互作用。一条单链寡核苷酸化学键合到直径为50纳米的单个银颗粒上,并且将Cy5标记的互补单链寡核苷酸与结合在银颗粒上的寡核苷酸杂交。荧光团与银颗粒之间的距离由长度为8纳米的刚性杂交DNA双链体维持。与不存在金属时的游离Cy5 - DNA分子相比,单个Cy5 - DNA - Ag颗粒的荧光强度增加了10倍以上,发射寿命降低了5倍。Cy5 - DNA - Ag颗粒寿命的降低使我们能够基于强度衰减解析两种物质的相关函数。与游离Cy5 - DNA相比,Cy5 - DNA - Ag颗粒亮度的增加导致Cy5 - DNA - Ag对混合物相关函数的贡献增加。这些结果表明,金属颗粒对荧光团的影响可用于在存在大量较暗物质的情况下检测金属结合物质的小部分群体。我们的结果还表明,这些与银颗粒缀合的明亮荧光团可作为荧光探针用于高背景生物样品中的临床检测。