Extracellular vesicles (EVs) have the potential to be
utilized
as disease-specific biomarkers. Although strategies for on-chip isolation
and detection of EVs have recently been developed, they need preprocessing
of clinical samples and are not accurate enough for disease diagnosis
just judging by EVs concentration. Here, we designed an integrated
microfluidic device named a plasma separation and EV detection (PS-ED)
chip for plasma separation, quantification, and high-throughput protein
analysis of EVs directly from clinical whole blood samples. The device
included two modules (PS and ED module): the PS module was a six-loop
microchannel for rapid separation of plasma from clinical whole blood
samples under inertial force; the amount of EVs in the separated plasma
kept the same value as in the initial blood samples. The module reduced
the mechanical damage to the blood cells and thus reduced the interference
of debris or cellular contents from damaged cells during EVs detection;
the ED module contained four S-channels for quantification and high-throughput
protein analysis of EVs; a wide detection range from 2.5 × 102 to 2.5 × 108 particles/μL with a detection
limit of 95 particles/μL was obtained. Through simultanously
monitoring three proteins (CD81, CD24, and EpCAM) of EVs, the cancer
type can be accurately confirmed. Furthermore, clinical blood sample
analysis verified that the proposed device could be used for accurate
diagnosis and therapy monitoring of ovarian cancer.
In this paper, we present a novel impedance microcytometer integrated with inertial focusing and liquid electrode techniques for high-throughput cell counting and discrimination. The inertial prefocusing unit orders cells into a determinate train to reduce the possibility of cell adhesions and ensure that only one cell passes through detection region at a time, which improves the accuracy of downstream detection. The liquid electrodes are constructed by inserting Ag/AgCl wires into the electrode chambers filled with flowing highly conductive electrolyte solutions, which have a high detection sensitivity while requiring a simple fabrication process. The effects of main sample flow rate, feed flow rate in electrode chambers, and feed solution type on measured impedance signals are experimentally explored. On the basis of the optimized system, we establish a linear relationship between the amplitude of impedance peaks and the volume of size-calibrated particles and achieve a high detection throughput of ∼5000 cells/s. Finally, using the calibrated microcytometer, we further investigate the size distributions of human breast tumor cells (MCF-7 cells) and leukocytes (white blood cells (WBCs)) and set a threshold amplitude to successfully distinguish the MCF-7 cells spiked in WBCs. Our impedance microcytometer may provide a potential tool for label-free cell enumeration and identification.
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