Department of Chemistry and Biochemistry, University of Delaware, Newark, DE 19716, USA.
Department of Biological Sciences, University of Delaware, Newark, DE 19716, USA; Department of Medical and Molecular Sciences, University of Delaware, Newark, DE 19716, USA.
Cell Rep Methods. 2024 Jun 17;4(6):100796. doi: 10.1016/j.crmeth.2024.100796. Epub 2024 Jun 11.
We present an efficient, effective, and economical approach, named E3technology, for proteomics sample preparation. By immobilizing silica microparticles into the polytetrafluoroethylene matrix, we develop a robust membrane medium, which could serve as a reliable platform to generate proteomics-friendly samples in a rapid and low-cost fashion. We benchmark its performance using different formats and demonstrate them with a variety of sample types of varied complexity, quantity, and volume. Our data suggest that E3technology provides proteome-wide identification and quantitation performance equivalent or superior to many existing methods. We further propose an enhanced single-vessel approach, named E4technology, which performs on-filter in-cell digestion with minimal sample loss and high sensitivity, enabling low-input and low-cell proteomics. Lastly, we utilized the above technologies to investigate RNA-binding proteins and profile the intact bacterial cell proteome.
我们提出了一种高效、经济且实用的蛋白质组学样品制备方法，命名为 E3 技术。通过将硅胶微球固定到聚四氟乙烯基质中，我们开发出一种坚固的膜介质，可作为一种可靠的平台，以快速且低成本的方式生成适合蛋白质组学的样品。我们使用不同的格式对其性能进行了基准测试，并使用各种复杂程度、数量和体积的不同类型的样品对其进行了演示。我们的数据表明，E3 技术提供的蛋白质组全谱鉴定和定量性能与许多现有方法相当或更优。我们进一步提出了一种增强型单容器方法，命名为 E4 技术，它可在过滤的同时进行细胞内消化，样品损失最小，灵敏度高，适用于低投入量和低细胞量的蛋白质组学研究。最后，我们利用上述技术研究 RNA 结合蛋白并描绘完整细菌细胞蛋白质组图谱。
