HLS-Scanning small laser beam flow cytometry:New concepts for microparticle analysis

 描述（由申请人提供）：流式细胞术是生物和医学科学的核心技术，近50年来一直由Mack Fulwyler于1965年发起，并由Herzenberg实验室推动进入免疫学领域以及许多其他领域。然而，核心原则在这段时间里几乎没有变化。虽然该技术的极限已经从1种发展到目前的最大18种荧光颜色和两种散射信号，但大多数仪器的工作范围都在该技术范围的中下层。该技术的基本原理在此期间保持不变，并且仅在每个细胞通过检测点时基于每个细胞测量总的蜂窝信号。这提供了丰富的数据源，但所收集的每个参数的内容很少。对于收集的每个变量，每个单元格都有一个值用于整个单元格。与共焦显微镜等技术的对比是清楚的。后者产生巨大的数据内容，但主要是基于更少的细胞，无法对悬浮细胞进行操作-流式细胞术的关键价值。使用细胞特性分离细胞以获得群体信息在成像中是困难的，并且物理分离实际上是不可能的。高容量筛选系统确实可以收集大量细胞的大量数据，但对这些细胞群的操作不如多参数流式细胞术复杂。本发明产生高含量和高细胞数据流，但重要的是，由于扫描技术，小颗粒的分辨率显著增加。我们的方法是由多年来在最高分辨率下对磁盘阅读和写技术的前沿开发而产生的。例如，DVD盘具有10 nm范围内的凹坑。通过将其中一些想法转移到使用蓝光中使用的类似尺寸的激光直径的微流体系统中，我们开辟了一个全新的机会，并将其定义为微扫描成像共聚焦流式细胞术。其结果是在以大约1 m/s的速度流动的细胞上产生了非常丰富的数据内容，扫描速率为1 MHz，以实现非常高的分辨率数据。现在存在一个主要的机会来重写从血细胞收集的信息，特别是微粒，这些微粒太小而不能被当前的流式细胞仪分析。当实现时，所提出的技术将能够收集许多荧光通道以及微小颗粒的详细形态数据，从而为全血或血浆以及其他生物样品中的先进微粒分析开辟新的篇章。

 DESCRIPTION (provided by applicant): Flow cytometry has been a core technology in biological and medical sciences for nearly 50 years having been initiated by Mack Fulwyler in 1965 and driven by the Herzenberg laboratory into the field of immunology as well as many other fields. However the core principles have changed little over that time. While the limits of the technology has advanced from 1 to a current maximum of 18 fluorescent colors and two scatter signals, the greater majority of instruments operate somewhere in the lower-middle of this technology range. The fundamental principles of the technology have remained the same over this time and only measures total cellular signals based on each cell as it passes the detection point. This provides a rich data source, but with low content with regard to each parameter collected. Each cell has a single value for the entire cell for each variable collected. The contrast with a technology such as confocal microscopy is clear. The latter produces tremendous data content, but mostly based on a fewer cells with inability to operate on suspended cells-the key value of flow cytometry. Separating cells using cellular properties to obtain population information is difficult in imaging, and physical separation is virtually impossible. It is true that high content screening systems can collect a lot of data on a lot of cells, but the manipulation of those populations is not as sophisticated as multiparameter flow cytometry. The present innovation produces both high content and high cell data streams but importantly with significant increase in resolution of small particles because of the scanning technology. Our approach is generated by many years of leading edge development of disk reading and writing technologies at the highest possible resolution. DVD discs for example have pits in the 10 nm range. By transferring some of these ideas onto a microfluidic system using laser diameters at similar sizes used in Blu-ray we have opened up an entirely new opportunity and have defined this as micro-scanning imaging confocal flow cytometry. The result is a tremendously rich data content on cells flowing at approximately 1 m/s with a scan rate of 1MHz to achieve very high resolution data. A major opportunity now exists to rewrite the information collected from blood cells and in particular microparticles which are too small to be analyzed by current flow cytometers. When realized, the proposed technology will be able to collect many channels of fluorescence as well as detailed morphological data on tiny particles opening up a new chapter in advanced micro particle analysis in whole blood or plasma as well as other biological samples.