Exosomes are secreted by most cell types and circulate in body fluids. Recent studies have revealed that exosomes play a significant role in intercellular communication and are closely associated with the pathogenesis of disease. Therefore, exosomes are considered promising biomarkers for disease diagnosis. However, exosomes are always mixed with other components of body fluids. Consequently, separation methods for exosomes that allow high-purity and high-throughput separation with a high recovery rate and detection techniques for exosomes that are rapid, highly sensitive, highly specific, and have a low detection limit are indispensable for diagnostic applications. For decades, many exosome separation and detection techniques have been developed to achieve the aforementioned goals. However, in most cases, these two techniques are performed separately, which increases operation complexity, time consumption, and cost. The emergence of microfluidics offers a promising way to integrate exosome separation and detection functions into a single chip. Herein, an overview of conventional and microfluidics-based techniques for exosome separation and detection is presented. Moreover, the advantages and drawbacks of these techniques are compared.

Progress in Microfluidics-Based Exosome Separation and Detection Technologies for Diagnostic Applications.

Several biomedical analyses are performed on particular types of cells present in body samples or using functionalized microparticles. Success in such analyses depends on the ability to separate or isolate the target cells or microparticles from the rest of the sample. In conventional procedures, multiple pieces of equipment, such as centrifuges, magnets, and macroscale filters, are used for such purposes, which are time-consuming, associated with human error, and require several operational steps. In the past two decades, there has been a tendency to develop microfluidic techniques, so-called lab-on-a-chip, to miniaturize and automate these procedures. The processes used for the separation and isolation of the cells and microparticles are scaled down into a small microfluidic chip, requiring very small amounts of sample. Differences in the physical and biological properties of the target cells from the other components present in the sample are the key to the development of such microfluidic techniques. These techniques are categorized as filtration-, hydrodynamic-, dielectrophoretic-, acoustic- and magnetic-based methods. Here we review the microfluidic techniques developed for sorting, separation, and isolation of cells and microparticles for biomedical applications. The mechanisms behind such techniques are thoroughly explained and the applications in which these techniques have been adopted are reviewed.

A review of sorting, separation and isolation of cells and microbeads for biomedical applications: microfluidic approaches.

Dielectric particles in a non-uniform electric field are subject to a force caused by a phenomenon called dielectrophoresis (DEP). DEP is a commonly used technique in microfluidics for particle or cell separation. In comparison with other separation methods, DEP has the unique advantage of being label-free, fast, and accurate. It has been widely applied in microfluidics for bio-molecular diagnostics and medical and polymer research. This review introduces the basic theory of DEP, its advantages compared with other separation methods, and its applications in recent years, in particular, focusing on the different electrode types integrated into microfluidic chips, fabrication techniques, and operation principles.

https://www.mdpi.com/2072-666X/10/6/423/pdf

DEP-on-a-Chip: Dielectrophoresis Applied to Microfluidic Platforms.

Detection of circulating tumour cells (CTCs) plays a crucial role in the diagnosis of tumour metastasis and evaluation of therapeutic effect on prognosis. The low number of the CTCs, however, makes their isolation from the peripheral blood very difficult. In this paper, a microfluidic device has been developed, which consists of a pair of straight interdigital transducers (IDTs) and focused interdigital transducers (FIDTs), to separate tumour cells from red blood cells (RBCs) in the blood stream by surface acoustic waves (SAWs). The standing SAWs (SSAW) generated by the straight IDTs enabled placing the CTCs and RBCs at the pressure nodes while the FIDTs generated the focused travelling pulsed SAWs (TSAWs) to push the CTCs away from the blood cells to achieve the isolation of CTCs. Using this mechanism, this device was capable of concentrating and separating 2 μm and 5 μm polystyrene particles and about 94.2% of particles with the diameter of 5 μm could be collected at the outlet. Moreover, 90% ± 2.4% of U87 glioma cells could be isolated from the RBCs when the cycle number of the TSAWs was 720 k. In the enrichment process, this microfluidic device totally relied on the SSAW without any additional assistance of the flow focusing. For the separation, unlike the bidirectional acoustic radiation force in SSAW, the unidirectional force generated by TSAWs consistently pushed the particles along the wave propagation, improving the sorting performance. This multi-stage device may provide an alternative method for sorting CTCs in a reliable and flexible manner.

Sorting of tumour cells in a microfluidic device by multi-stage surface acoustic waves

Review: Microbial analysis in dielectrophoretic microfluidic systems

Screen-printed microfluidic dielectrophoresis chip for cell separation

The invention discloses a method for manufacturing microfluidic dielectrophoresis chips through the screen printing technology The method includes the following steps that first, a microelectrode pattern and a microchannel pattern are drawn, and a microelectrode screen printing plate and a microchannel screen printing plate are manufactured; second, through the microelectrode screen printing plate, conductive ink is printed on printed material, and a microelectrode is manufactured; third, through the microchannel screen printing plate, insulating ink capable of solidification is printed on the printed material (obtained in the second step) on which the microelectrode is printed with a target alignment method, and a microchannel is manufactured; fourth, the portion above the microchannel is covered with a sealing-connection cover sheet and solidified so that the sealing-connection cover sheet can be connected with the microchannel in a sealed mode The method effectively solves the problems that a traditional microfluidic dielectrophoresis chip manufacturing method is long in manufacturing period, low in efficiency and not suitable for industrial production on a large scale

Method for manufacturing microfluidic dielectrophoresis chips through screen printing technology

A novel amperometric immunosensor was fabricated using an electropolymerized conducting polymer composite of pyrrole and 4-(3-pyrrolyl) butyric acid as the probe immobilization matrix. The biomolecule was immobilized via covalent bonding on the polymer surface with a high density of carboxyl groups. Instead of directly attaching the probe to the polymer surface, the streptavidin-biotin affinity complex was introduced to improve the bioactivity, orientation, and the amount of immobilized material. Polyvinyl alcohol was employed to improve the hydrophilicity of the polymer surface and further decrease the nonspecific protein adsorption. As a result, high sensitivity and specificity were achieved with a wide linear dynamic range. Using mouse immunoglobulin G as a model protein, the immunosensor exhibited a linear relationship between 7 ng/mL and 20 µg/mL with a limit of detection of 7 ng/mL.

A Novel Electrochemical Immunosensor Incorporating a Pyrrole/4-(3-Pyrrolyl) Butyric Acid Conducting Polymer

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Screen-printed microfluidic dielectrophoresis chip for cell separation

Method for manufacturing microfluidic dielectrophoresis chips through screen printing technology

A Novel Electrochemical Immunosensor Incorporating a Pyrrole/4-(3-Pyrrolyl) Butyric Acid Conducting Polymer