Stanisheuski Stanislau, Ebrahimi Arpa, Vaidya Kavi Aashish, Jang Hyo Sang, Yang Liping, Eddins Alex Jordan, Marean-Reardon Carrie, Franco Maria Clara, Maier Claudia Susanne
Department of Chemistry, Oregon State University, Corvallis, OR, United States.
Department of Biochemistry and Biophysics, College of Science, Oregon State University, Corvallis, OR, United States.
Front Chem. 2024 Aug 21;12:1428547. doi: 10.3389/fchem.2024.1428547. eCollection 2024.
In this study, we adapted an HP D100 Single Cell Dispenser - a novel low-cost thermal inkjet (TIJ) platform with impedance-based single cell detection - for dispensing of individual cells and one-pot sample preparation. We repeatedly achieved label-free identification of up to 1,300 proteins from a single cell in a single run using an Orbitrap Fusion Lumos Mass Spectrometer coupled to either an Acquity UPLC M-class system or a Vanquish Neo UHPLC system. The developed sample processing workflow is highly reproducible, robust, and applicable to standardized 384- and 1536-well microplates, as well as glass LC vials. We demonstrate the applicability of the method for proteomics of single cells from multiple cell lines, mixed cell suspensions, and glioblastoma tumor spheroids. As additional proof of robustness, we monitored the results of genetic manipulations and the expression of engineered proteins in individual cells. Our cost-effective and robust single-cell proteomics workflow can be transferred to other labs interested in studying cells at the individual cell level.
在本研究中，我们采用了惠普D100单细胞分配仪——一种基于阻抗单细胞检测的新型低成本热喷墨（TIJ）平台——用于单个细胞的分配和单管样品制备。我们使用与Acquity UPLC M级系统或Vanquish Neo超高效液相色谱系统联用的Orbitrap Fusion Lumos质谱仪，单次运行可从单个细胞中反复实现多达1300种蛋白质的无标记鉴定。所开发的样品处理工作流程具有高度可重复性、稳健性，适用于标准化的384孔和1536孔微孔板以及玻璃液相色谱小瓶。我们展示了该方法在多种细胞系、混合细胞悬液和胶质母细胞瘤肿瘤球体单细胞蛋白质组学中的适用性。作为稳健性的额外证据，我们监测了单个细胞中基因操作的结果和工程蛋白的表达。我们这种经济高效且稳健的单细胞蛋白质组学工作流程可转移到其他有兴趣在单个细胞水平研究细胞的实验室。
