A submersible imaging-in-flow instrument to monitor nano- and microplankton

拟议的工作建立在“FlowCytobot”的基础上，这是一种由pi开发的潜水流式细胞仪，用于观察10-100米范围内的浮游植物和微型浮游动物。这个间隙包含可以在沿海水域繁殖的浮游植物（包括许多硅藻和鞭毛藻），以及微型植物动物，如原生动物，它们对桡足类和幼鱼的饮食至关重要，因此我们对这个大小范围内的生物进行监测是很重要的。本提案的目标是完成潜水成像流式细胞仪的开发和测试，旨在表征这些大型浮游植物和微型浮游动物生物。这种新仪器是FlowCytobot的改进版本，它结合了流式细胞术和视频技术，捕捉生物体的高分辨率（1米）图像进行识别，并测量与每张图像相关的叶绿素荧光。图像将由基于支持向量机方法的软件进行分类，而叶绿素荧光的测量将使我们能够在包含两种成员的分类群（如鞭毛藻）中区分异养和光养细胞。大型浮游植物细胞中叶绿素荧光的定量也将使大量叶绿素数据中的模式得到更好的解释。Imaging FlowCytobot的工作版本的构建已接近完成。新仪器保留了原有FlowCytobot的防污和性能监测功能；它可以自主操作，但可以远程编程并连续采样；数据将通过以太网和光纤电缆传输到远程计算机进行分析。接下来的开发步骤包括在WHOI和MVCO进行现场测试，根据测试结果优化仪器，并改进分析和控制软件。

The proposed work builds on the "FlowCytobot", a submersible flow cytometer developed by the PIs to observe phytoplankton and microzooplankton in the size range 10-100 m. This gap contains phytoplankton (including many diatoms and dinoflagellates) that can bloom in coastal waters, and microzooplanton such as protozoa that are critical to the diets of copepods and larval fish, so it is important that we monitor the organisms in this size range. The goal of this proposal, is to complete development and testing of a submersible imaging flow cytometer designed to characterize these large phytoplankton and microzooplanktonic organisms. The new instrument is a modified version of FlowCytobot that uses a combination of flow cytometric and video technology to capture high resolution (1 m) images of organisms for identification and to measure chlorophyll fluorescence associated with each image. Images will be classified by software based on a support vector machine approach, while the measurements of chlorophyll fluorescence will allow us to discriminate heterotrophic from phototrophic cells in taxa (such as dinoflagellates) that contain both kinds of members. Quantification of chlorophyll fluorescence in large phytoplankton cells will also enable patterns in bulk chlorophyll data to be better interpreted.Construction of a working version of Imaging FlowCytobot is nearly completed. The new instrument retains the anti-fouling and performance-monitoring capabilities of the original FlowCytobot; it operates autonomously but is remotely programmable and samples continuously; data will be transferred via Ethernet and fiber optic cables to a remote computer for analysis. The next steps in development involve field testing at WHOI and MVCO, optimizing the instrument on the basis of results from this testing, and refining the analysis and control software.