TY - JOUR
T1 - Temperature-Responsive Agarose-Based Digital Microfluidics
T2 - An In-Chip Sample Preparation To Mass Spectrometry Analysis for Trace Cells and Single-Cell Proteomics
AU - Yan, Ling
AU - Fang, Jiacheng
AU - Zhang, Huimin
AU - Lin, Yali
AU - Li, Dan
AU - Zhou, Yingyan
AU - Zhu, Lin
AU - Zhu, Zhi
AU - Yang, Pengyuan
AU - Cai, Zongwei
N1 - This study was supported by the Tier 1 Research Start-up Grants from the Research Committee of Hong Kong Baptist University (162874) and the donation from the Kwok Chung Bo Fun Charitable Fund for the establishment of the Kwok Yat Wai Endowed Chair of Environmental and Biological Analysis. L.Y. and L.Z. acknowledge the facility support from the Advanced Life Sciences and Mass Spectrometry Laboratory (LSMS) of Hong Kong Baptist University.
Publisher Copyright:
© 2026 American Chemical Society
PY - 2026/1/27
Y1 - 2026/1/27
N2 - Mass spectrometry (MS)-based single- and trace-cell proteomics provides critical insights into cellular phenotypes, but widespread use is limited by the cost and complexity of advanced MS systems. We present a cost-effective, accessible workflow compatible with standard MS platforms and scalable for multiomics. The temperature-responsive agarose-based digital microfluidics (TRA-DMF) platform enables one-step sample processing, including lysis, reduction/alkylation, and digestion, in a parallel four-channel format. Unlike conventional droplet-based microfluidics or fluorescence-activated cell sorting (FACS) approaches, our DMF system ensures real-time visualization and confirmation of single-cell capture and 98.3% sample recovery, minimizing losses through nonpipetting transfer. The TRA-DMF system also overcomes the MS-incompatibility of oil-phase microfluidics, allowing high-efficiency droplet transfer to MS vials. The entire TRA-DMF for single cell proteomics (TRA-DMF-SCP) workflow is completed in ∼3 h (including single-cell capture), with seamless sample introduction into standard LC-MS/MS systems. Using a regular benchtop MS instrument Orbitrap Fusion, we identified over 4000 protein groups (PGs) from 50 293T cells with excellent reproducibility and robustness. This system offers a practical and scalable solution for trace- and single-cell proteomics and holds strong potential for integration into routine multiomics workflows.
AB - Mass spectrometry (MS)-based single- and trace-cell proteomics provides critical insights into cellular phenotypes, but widespread use is limited by the cost and complexity of advanced MS systems. We present a cost-effective, accessible workflow compatible with standard MS platforms and scalable for multiomics. The temperature-responsive agarose-based digital microfluidics (TRA-DMF) platform enables one-step sample processing, including lysis, reduction/alkylation, and digestion, in a parallel four-channel format. Unlike conventional droplet-based microfluidics or fluorescence-activated cell sorting (FACS) approaches, our DMF system ensures real-time visualization and confirmation of single-cell capture and 98.3% sample recovery, minimizing losses through nonpipetting transfer. The TRA-DMF system also overcomes the MS-incompatibility of oil-phase microfluidics, allowing high-efficiency droplet transfer to MS vials. The entire TRA-DMF for single cell proteomics (TRA-DMF-SCP) workflow is completed in ∼3 h (including single-cell capture), with seamless sample introduction into standard LC-MS/MS systems. Using a regular benchtop MS instrument Orbitrap Fusion, we identified over 4000 protein groups (PGs) from 50 293T cells with excellent reproducibility and robustness. This system offers a practical and scalable solution for trace- and single-cell proteomics and holds strong potential for integration into routine multiomics workflows.
UR - https://www.scopus.com/pages/publications/105029528903
U2 - 10.1021/acs.analchem.5c06267
DO - 10.1021/acs.analchem.5c06267
M3 - Journal article
AN - SCOPUS:105029528903
SN - 0003-2700
VL - 98
SP - 2285
EP - 2296
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 3
ER -