Multiplex Profiling of Biomarker and Drug Uptake in Single Cells Using Microfluidic Flow Cytometry and Mass Spectrometry

Abstract

To perform multiplex profiling of single cells and eliminate the risk of potential sample loss caused by centrifugation, we developed a microfluidic flow cytometry and mass spectrometry system (μCytoMS) to evaluate the drug uptake and induced protein expression at the single cell level. It involves a microfluidic chip for the alignment and purification of single cells followed by detection with laser-induced fluorescence (LIF) and inductively coupled plasma mass spectrometry (ICP-MS). Biofunctionalized nanoprobes (Bi-oNPs), conjugating ∼3000 6-FAM-Sgc8 aptamers on a single gold nanoparticle (AuNP) (K d = 0.23 nM), were engineered to selectively bind with protein tyrosine kinase 7 (PTK7) on target cells. PTK7 expression induced by oxaliplatin (OXA) uptake was assayed with LIF, while ICP-MS measurement of 195 Pt revealed OXA uptake of the drug in individual cells, which provided further in-depth information about the drug in relation to PTK7 expression. At an ultralow flow of ∼0.043 dyn/cm 2 (20 μL/min), the chip facilitates the extremely fast focusing of BioNPs labeled single cells without the need for centrifugal purification. It ensures multiplex profiling of single cells at a throughput speed of 500 cells/min as compared to 40 cells/min in previous studies. Using a machine learning algorithm to initially profile drug uptake and marker expression in tumor cell lines, μCytoMS was able to perform in situ profiling of the PTK7 response to the OXA at single-cell resolution for tests done on clinical samples from 10 breast cancer patients. It offers great potential for multiplex single-cell phenotypic analysis and clinical diagnosis.

A recyclable PDMS microfluidic surface-enhanced Raman scattering Cu/AgNP chip for the analysis of sulfadiazine in aquatic products

“…Microfluidic chips facilitate the regulation of sample flow and mixing via microchannels and microvalves, enabling efficient sample processing and detection. 5,6 Additionally, by integrating various detection methods such as mass spectrometry analysis, 7 Raman spectroscopy, 8 fluorescence detection 9 and so on, microfluidic chips can be applied in fields such as biological analysis, 10 environmental monitoring, 11 and drug screening. 12 For example, by combining microfluidic chips with mass spectrometry, Zeng 13 developed a new type of nanoelectrospray emitter based on microfluidic manufacture, offering stable electrospray, reproducibility, and enhanced ionization efficiency for bioanalyses.…”

Stabilizing and Accelerating Secondary Flow in Ultralong Spiral Channel for High-Throughput Cell Manipulation

“…To enhance the throughput, the optimal solution is to expand the channel dimension. Spiral-cured channels are the most suitable for size design due to their extended channel length, rational space arrangement, and distinct Dean secondary flow. , However, designing a spiral channel to efficiently manipulate cells remains challenging due to the varying Dean secondary flow derived from the differences in the radius of each loop. Since Dean secondary flow is also sensitive to flow rates, it may take multiple attempts to ascertain the suitable flow rates. , The lack of general and effective means to regulate the Dean secondary flow has limited the high efficiency of spiral microchannels in cell manipulation.…”