Wu Chengxin, Wei Xing, Men Xue, Xu Yulong, Bai Junjie, Wang Yu, Zhou Lei, Yu Yong-Liang, Xu Zhang-Run, Chen Ming-Li, Wang Jian-Hua
Research Center for Analytical Sciences, Department of Chemistry, College of Sciences, Northeastern University, Shenyang, 110819, China.
State Key Laboratory of Applied Organic Chemistry, College of Chemistry and Chemical Engineering, Lanzhou University, Lanzhou, 730000, China.
Talanta. 2023 Jun 1;258:124424. doi: 10.1016/j.talanta.2023.124424. Epub 2023 Mar 6.
Flow cytometry is among the most powerful tools for single-cell analysis, while the high cost and mechanical complexity of the commercial instrumentation limit the applications in personalized single-cell analysis. For this issue, we hereby construct an open and low-cost flow cytometer. It is highly compact to integrate the functions of (1) single cell aligning by a lab-made modularized 3D hydrodynamic focusing device, and (2) fluorescence detection of the single cells by a confocal laser-induced fluorescence (LIF) detector. The ceiling cost of the entire hardware for the LIF detection unit and 3D focusing device is $ 3200 and $ 400 respectively. A sheath flow velocity of 150 μL/min produces a focused sample stream of 17.6 μm × 14.6 μm at sample flow of 2 μL/min, based on the LIF response frequency and the laser beam spot diameter. The assay performance of the flow cytometer was evaluated by characterizing fluorescent microparticles and acridine orange (AO) stained HepG2 cells, producing throughputs of 40.5/s and 6.2/s respectively. Favorable assay precision and accuracy were demonstrated by the agreement of frequency histogram with imaging analysis, and good Gaussian-like distributions of fluorescent microparticles and AO-stained HepG2 cells. Practically, the flow cytometer was successfully applied for the evaluation of ROS generation in single HepG2 cells.
流式细胞术是单细胞分析最强大的工具之一，然而商业仪器的高成本和机械复杂性限制了其在个性化单细胞分析中的应用。针对这一问题，我们特此构建了一种开放式低成本流式细胞仪。它高度紧凑，集成了以下功能：（1）通过实验室自制的模块化三维流体动力聚焦装置对单细胞进行对齐；（2）通过共聚焦激光诱导荧光（LIF）探测器对单细胞进行荧光检测。LIF检测单元和三维聚焦装置的整个硬件的最高成本分别为3200美元和400美元。基于LIF响应频率和激光束光斑直径，在样品流速为2μL/min时，鞘液流速为150μL/min可产生17.6μm×14.6μm的聚焦样品流。通过对荧光微球和吖啶橙（AO）染色的HepG2细胞进行表征，评估了流式细胞仪的检测性能，其通量分别为40.5个/秒和6.2个/秒。频率直方图与成像分析的一致性以及荧光微球和AO染色的HepG2细胞良好的类高斯分布证明了良好的检测精度和准确性。实际上，该流式细胞仪已成功应用于评估单个HepG2细胞中活性氧的产生。
