National Flow Cytometry Resource, Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA.
J Am Chem Soc. 2010 May 5;132(17):6081-90. doi: 10.1021/ja909850s.
Nanoparticle spectroscopic tags based on surface enhanced Raman scattering (SERS) are playing an increasingly important role in bioassay and imaging applications. The ability to rapidly characterize large populations of such tags spectroscopically in a high-throughput flow-based platform will open new areas for their application and provide new tools for advancing their development. We demonstrate here a high-resolution spectral flow cytometer capable of acquiring Raman spectra of individual SERS-tags at flow rates of hundreds of particles per second, while maintaining the spectral resolution required to make full use of the detailed information encoded in the Raman signature for advanced multiplexing needs. The approach allows multiple optical parameters to be acquired simultaneously over thousands of individual nanoparticle tags. Characteristics such as tag size, brightness, and spectral uniformity are correlated on a per-particle basis. The tags evaluated here display highly uniform spectral signatures, but with greater variability in brightness. Subpopulations in the SERS response, not apparent in ensemble measurements, are also shown to exist. Relating tag variability to synthesis parameters makes flow-based spectral characterization a powerful tool for advancing particle development through its ability to provide rapid feedback on strategies aimed at constraining desired tag properties. Evidence for single-tag signal saturation at high excitation power densities is also shown, suggesting a role for high-throughput investigation of fundamental properties of the SERS tags as well.
基于表面增强拉曼散射（SERS）的纳米粒子光谱标记物在生物测定和成像应用中发挥着越来越重要的作用。在高通量流基平台上快速对大量此类标记物进行光谱特征描述的能力将为它们的应用开辟新的领域，并为推进它们的发展提供新的工具。我们在这里展示了一种高分辨率光谱流动细胞仪，能够以每秒数百个颗粒的流速获取单个 SERS 标记物的拉曼光谱，同时保持充分利用拉曼特征中编码的详细信息所需的光谱分辨率，以满足高级复用的需求。该方法允许在数千个单个纳米粒子标记物上同时获取多个光学参数。基于每个粒子的基础上，对标记物的大小、亮度和光谱均匀性等特征进行相关分析。这里评估的标记物显示出高度均匀的光谱特征，但亮度变化较大。还显示出在集合测量中不明显的 SERS 响应的子群体的存在。将标记物的可变性与合成参数相关联，使基于流的光谱特性成为一种强大的工具，能够通过快速反馈来控制所需的标记物特性，从而推进粒子的发展。还表明在高激发功率密度下存在单个标记物信号饱和的证据，这表明也需要高通量研究 SERS 标记物的基本特性。
