Institute of Physics, Leiden University, Post Office Box 9504, 2300 RA Leiden, Netherlands.
Science. 2010 Oct 15;330(6002):353-6. doi: 10.1126/science.1195475.
So far, single-molecule imaging has predominantly relied on fluorescence detection. We imaged single nonfluorescent azo dye molecules in room-temperature glycerol by the refractive effect of the heat that they release in their environment upon intense illumination. This photothermal technique provides contrast for the absorbing objects only, irrespective of scattering by defects or roughness, with a signal-to-noise ratio of ~10 for a single molecule in an integration time of 300 milliseconds. In the absence of oxygen, virtually no bleaching event was observed, even after more than 10 minutes of illumination. In a solution saturated with oxygen, the average bleaching time was of the order of 1 minute. No blinking was observed in the absorption signal. On the basis of bleaching steps, we obtained an average absorption cross section of 4 angstroms(2) for a single chromophore.
到目前为止，单分子成像主要依赖于荧光检测。我们通过在强烈照明下它们在环境中释放的热量的折射效应，对单个非荧光偶氮染料分子在室温甘油中进行成像。这种光热技术仅为吸收物体提供对比度，而与缺陷或粗糙度的散射无关，在 300 毫秒的积分时间内，单个分子的信噪比约为 10。在没有氧气的情况下，即使在照明超过 10 分钟后，也几乎没有观察到漂白事件。在氧气饱和的溶液中，平均漂白时间约为 1 分钟。在吸收信号中没有观察到闪烁。根据漂白步骤，我们获得了单个生色团的平均吸收截面为 4 埃（2）。
