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Efficient cell manipulation is essential for numerous applications in bioanalysis and medical diagnosis. However, the lack of stability and strength in the secondary flow, coupled with the narrow range of practical throughput, severely restricts the diverse applications. Herein, we present an innovative inertial microfluidic device that employs a spiral channel for high-throughput cell manipulation. Our investigation demonstrates that the regulation of Dean-like secondary flow in the microchannel can be achieved through geometric confinement. Introducing ordered microstructures into the ultralong spiral channel (>90 cm) stabilizes and accelerates the secondary flow among different loops. Consequently, effective manipulation of blood cells within a wide cell throughput range (1.73 × 108 to 1.16 × 109 cells/min) and cancer cells across a broad throughput range (0.5 × 106 to 5 × 107 cells/min) can be achieved. In comparison to previously reported technologies, our engineering approach of stabilizing and accelerating secondary flow offers specific performance for cell manipulation under a wide range of high-throughput manner. This engineered spiral channel would be promising in biomedical analysis, especially when cells need to be focused efficiently on large-volume liquid samples.
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Dispositivos Laboratorio en un Chip , Humanos , Técnicas Analíticas Microfluídicas/instrumentación , Ensayos Analíticos de Alto RendimientoRESUMEN
Single-cell encapsulation in droplet microfluidics is commonly hindered by the tradeoff between cell suspension density and on-chip focusing performance. In this study, we introduce a novel droplet microfluidic chip to overcome this challenge. The chip comprises a double spiral focusing unit, a flow resistance-based sample enrichment module with fine-tunable outlets, and a crossflow droplet generation unit. Utilizing a low-density cell/bead suspension (2 × 106 objects/mL), cells/beads are focused into a near-equidistant linear arrangement within the double spiral microchannel. The excess water phase is diverted while cells/beads remain focused and sequentially encapsulated in individual droplets. Focusing performance was assessed through numerical simulations and experiments at three flow rates (40, 60, 80 µL/min), demonstrating successful focusing at 40 and 80 µL/min for beads and cells, respectively. In addition, both simulation and experimental results revealed that the flow resistance at the sample enrichment module is adjustable by punching different outlets, allowing over 50% of the aqueous phase to be removed. YOLOv8n-based droplet detection algorithms realized the counting of cells/beads in droplets, statistically demonstrating single-cell and bead encapsulation rates of 72.2% and 79.2%, respectively. All the results indicate that this on-chip sample enrichment approach can be further developed and employed as a critical component in single-cell encapsulation in water-in-oil droplets.
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Inertial microfluidics has demonstrated its ability to focus particles in a passive and straightforward manner. However, achieving flow-rate- and particle-size-insensitive focusing in large-dimension channels with a simple design remains challenging. In this study, we developed a spiral microfluidic with a large-dimension channel to achieve inertial focusing. By designing a unique "big buffering area" and a "small buffering area" in the spiral microchannel, we observed the stabilization and acceleration of secondary flow. Our optimized design allowed for efficient (>99.9%) focusing of 15 µm particles within a wide range of flow rates (0.5-4.5 mL/min) during a long operation duration (0-60 min). Additionally, we achieved effective (>95%) focusing of different-sized particles (7, 10, 15, and 30 µm) and three types of tumor cells (K562, HeLa, and MCF-7) near the inner wall of the 1 mm wide outlet when applying different flow rates (1-3 mL/min). Finally, successful 3D cell focusing was achieved within an optimized device, with the cells positioned at a distance of 50 µm from the wall. Our strategy of stabilizing and accelerating Dean-like secondary flow through the unique configuration of a "big buffering area" and a "small buffering area" proved to be highly effective in achieving inertial focusing that is insensitive to the flow rate and particle size, particularly in large-dimension channels. Consequently, it shows great potential for use in hand-operated microfluidic tools for flow cytometry.
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Técnicas Analíticas Microfluídicas , Microfluídica , Humanos , Microfluídica/métodos , Tamaño de la Partícula , Citometría de Flujo/métodos , AceleraciónRESUMEN
Microfluidic technologies enabling the control of secondary flow are essential for the successful separation of blood cells, a process that is beneficial for a wide range of medical research and clinical diagnostics. Herein, we introduce a dimension-confined microfluidic device featuring a double-spiral channel designed to regulate secondary flows, thereby enabling high-throughput isolation of blood for plasma extraction. By integrating a sequence of micro-obstacles within the double-spiral microchannels, the stable and enhanced Dean-like secondary flow across each loop can be generated. This setup consequently prompts particles of varying diameters (3, 7, 10, and 15 µm) to form different focusing states. Crucially, this system is capable of effectively separating blood cells of different sizes with a cell throughput of (2.63-3.36) × 108 cells/min. The concentration of blood cells in outlet 2 increased 3-fold, from 1.46 × 108 to 4.37 × 108, while the number of cells, including platelets, exported from outlets 1 and 3 decreased by a factor of 608. The engineering approach manipulating secondary flow for plasma extraction points to simplicity in fabrication, ease of operation, insensitivity to cell size, high throughput, and separation efficiency, which has potential utility in propelling the development of miniaturized diagnostic devices in the field of biomedical science.
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Técnicas Analíticas Microfluídicas , Microfluídica , Células Sanguíneas , Plasma , PlaquetasRESUMEN
Although microfluidic devices have made remarkable strides in blood cell separation, there is still a need for further development and improvement in this area. Herein, we present a novel ultralow aspect ratio (H/W = 1:36) spiral channel microfluidic device with ordered micro-obstacles for sheathless and flow-rate-insensitive blood cell separation. By introducing ordered micro-obstacles into the spiral microchannels, reduced magnitude fluctuations in secondary flow across different loops can be obtained through geometric confinement. As a result, the unique Dean-like secondary flow can effectively enhance the separation efficiency of particles in different sizes ranging from 3 to 15 µm. Compared to most existing microfluidic devices, our system offers several advantages of easy manufacturing, convenient operation, long-term stability, highly efficient performance (up to 99.70% rejection efficiency, including platelets), and most importantly, insensitivity to cell sizes as well as flow rates (allowing for efficient separation of different-sized blood cells in a wide flow rate from 1.00 to 2.50 mL/min). The unique characteristics, such as ultralow aspect ratio, sequential micro-obstacles, and controlled secondary flow, make our device a promising solution for practical plasma extraction in biomedical research and clinical applications.
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Técnicas Analíticas Microfluídicas , Plasma , Células Sanguíneas , Plaquetas , AceleraciónRESUMEN
A aerobic, gram-negative, rod-shaped and polar-flagellum bacterial strain, designated as FYR11-62T, was isolated from the estuary of the Fenhe River into the Yellow River in Shanxi Province, China. The isolate was able to grow at 4-37 °C (optimum, 25 °C), pH 5.5-9.5 (optimum, pH 7.5) and in the presence of 0-7.0% (w/v) NaCl (optimum, 1.0% NaCl). Phylogenetic analyses based on 16S rRNA genes and 1597 single-copy orthologous clusters indicated that strain FYR11-62T affiliated with the genus Shewanella and shared the highest 16S rRNA gene sequence similarity to Shewanella aestuarii SC18T (98.3%) and Shewanella gaetbuli TF-27T (97.3%), respectively. The major fatty acids were summed feature 3 (C16:1 ω7c and/or C16:1 ω6c), C16:0 and iso-C15:0. The major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. The main quinones were Q-7 and Q-8. The genomic DNA G + C content was 41.6%. Gene annotation showed that strain FYR11-62T possessed 30 antibiotic resistance genes, implying its multiple antidrug resistance. The average nucleotide identity and digital DNA-DNA hybridization values between strain FYR11-62T and its closely related species were all below the thresholds for species delineation. The phylogenetic position together with the results of the analysis of morphological, physiological and genomic features support the classification of strain FYR11-62T (= MCCC 1K07242T = KCTC 92244T) as a novel species of the genus Shewanella, for which the name Shewanella subflava sp. nov. is proposed.
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Ríos , Shewanella , Ríos/microbiología , Filogenia , Estuarios , ARN Ribosómico 16S/genética , Cloruro de Sodio , Análisis de Secuencia de ADN , Ácidos Grasos/análisis , Técnicas de Tipificación Bacteriana , ADN Bacteriano/genética , ADN Bacteriano/química , Fosfolípidos/análisisRESUMEN
Isolation and manipulation of single cells play a crucial role in drug screening. However, previously reported single-cell drug screening lacked multiple-dose concentration gradient studies, which limits their ability to predict drug performance accurately. To solve this problem, we constructed a multiconcentration gradient generator in which a Tai Chi-spiral mixer can accelerate solution mixing in a short time and produce a linear concentration gradient. Later, a gradient generator combined with a single-cell capture array was adopted to investigate the effects of single or combined doses of 5-fluorouracil and cisplatin on human hepatoma cells and human breast carcinoma cells (at the single-cell level). The results showed that both drugs were effective in inhibiting the growth of cancer cells, and the combination was more effective for human hepatoma cells. In addition, the relationship between the biomechanical heterogeneity (e.g., deformability and size) of tumor cells and potential drug resistance at the single-cell level was investigated, indicating that small and/or deformable cells were more resistant than large and/or less deformable cells. The device provides a simple and reliable platform for studying the optimal dosage of different drug candidates at the single-cell level and effectively screening single-agent chemotherapy regimens and combination therapies.
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Inertial microfluidics enables fluid and particle manipulation for biomedical and clinical applications. Herein, we developed a simple semicircular microchannel with an ultra-low aspect ratio to interrogate the unique formations of the helical vortex and Dean vortex by introducing order micro-obstacles. The purposeful and powerful regulation of dimensional confinement in the microchannel achieved significantly improved fluid mixing effects and fluid and particle manipulation in a high-throughput, highly efficient and easy-to-use way. Together, the results offer insights into the geometry-induced multi-vortex mechanism, which may contribute to simple, passive, continuous operations for biochemical and clinical applications, such as the detection and isolation of circulating tumor cells for cancer diagnostics.
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The field of inertial microfluidics has been significantly advanced in terms of application to fluid manipulation for biological analysis, materials synthesis, and chemical process control. Because of their superior benefits such as high-throughput, simplicity, and accurate manipulation, inertial microfluidics designs incorporating channel geometries generating Dean vortexes and helical vortexes have been studied extensively. However, existing technologies have not been studied by designing low-aspect-ratio microchannels to produce multi-vortexes. In this study, an inertial microfluidic device was developed, allowing the generation and regulation of the Dean vortex and helical vortex through the introduction of micro-obstacles in a semicircular microchannel with ultra-low aspect ratio. Multi-vortex formations in the vertical and horizontal planes of four dimension-confined curved channels were analyzed at different flow rates. Moreover, the regulation mechanisms of the multi-vortex were studied systematically by altering the micro-obstacle length and channel height. Through numerical simulation, the regulation of dimensional confinement in the microchannel is verified to induce the Dean vortex and helical vortex with different magnitudes and distributions. The results provide insights into the geometry-induced secondary flow mechanism, which can inspire simple and easily built planar 2D microchannel systems with low-aspect-ratio design with application in fluid manipulations for chemical engineering and bioengineering.
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Malva vein clearing virus (MVCV) is a member of the Potyvirus species, and has a negative impact on the aesthetic development of Alcea rosea. It was first reported in Germany in 1957, but its complete genome sequence data are still scarce. In the present work, A. rosea leaves with vein-clearing and mosaic symptoms were sampled and analyzed with small RNA deep sequencing. By denovo assembly the raw sequences of virus-derived small interfering RNAs (vsiRs) and whole genome amplification of malva vein cleaning virus SX strain (MVCV-SX) by specific primers targeting identified contig gaps, the full-length genome sequences (9,645 nucleotides) of MVCV-SX were characterized, constituting of an open reading frame that is long enough to encode 3,096 amino acids. Phylogenetic analysis showed that MVCV-SX was clustered with euphorbia ringspot virus and yam mosaic virus. Further analyses of the vsiR profiles revealed that the most abundant MVCV-vsiRs were between 21 and 22 nucleotides in length and a strong bias was found for "A" and "U" at the 5'-terminal residue. The results of polarity assessment indicated that the amount of sense strand was almost equal to that of the antisense strand in MVCV-vsiRs, and the main hot-spot region in MVCV-SX genome was found at cylindrical inclusion. In conclusion, our findings could provide new insights into the RNA silencing-mediated host defence mechanism in A. rosea infected with MVCV-SX, and offer a basis for the prevention and treatment of this virus disease.
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With the continuous development of cancer therapy, conventional animal models have exposed a series of shortcomings such as ethical issues, being time consuming and having an expensive cost. As an alternative method, microfluidic devices have shown advantages in drug screening, which can effectively shorten experimental time, reduce costs, improve efficiency, and achieve a large-scale, high-throughput and accurate analysis. However, most of these microfluidic technologies are established for narrow-range drug-concentration screening based on sensitive but limited flow rates. More simple, easy-to operate and wide-ranging concentration-gradient constructions for studying tumor cell-drug interactions in real-time have remained largely out of reach. Here, we proposed a simple and compact device that can quickly construct efficient and reliable drug-concentration gradients with a wide range of flow rates. The dynamic study of concentration-gradient formation based on successive spiral mixer regulations was investigated systematically and quantitatively. Accurate, stable, and controllable dual drug-concentration gradients were produced to evaluate simultaneously the efficacy of the anticancer drug against two tumor cell lines (human breast adenocarcinoma cells and human cervical carcinoma cells). Results showed that paclitaxel had dose-dependent effects on the two tumor cell lines under the same conditions, respectively. We expect this device to contribute to the development of microfluidic chips as a portable and economical product in terms of the potential of concentration gradient-related biochemical research.
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Research and development of innovative targeted therapies is a great challenge in the fight against cancer. Although many treatment methods are currently available, there is no simple and effective system for promptly conducting anti-cancer drug screening and dose-response evaluation of the cancer patients to the drug. Herein, we developed an easy and compact flow rate independent microfluidic chip that can rapidly construct three concentration gradients of multiple solutes based on Dean flow under a wide range of flow rates. Chemical gradient dynamics were investigated systematically and quantitatively. Three stable, accurate, and controllable drug gradients were generated to evaluate treatments of two tumor cell lines (MCF-7 and HepG2). Results showed the dose- and time-dependent antitumor effects of the drugs, indicating the suitability of the proposed system to evaluate the individual actions and interactions of the anti-cancer drugs (doxorubicin and cisplatin) on one tumor cell line under the same conditions. In addition, cell viability in the microfluidic chip under gradient conditions showed a linear relationship to the viability of the traditional culture experiment. In summary, our microfluidic device can be used to develop insensitive techniques to operational conditions for simultaneously establishing multi-drug concentration gradients, which has the potential to promote the development of specific drug screening tools for targeting multiple vulnerabilities of tumor cells and evaluating the most effective personalized treatment technique.
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Ensayos de Selección de Medicamentos Antitumorales/métodos , Dispositivos Laboratorio en un Chip , Antineoplásicos/farmacología , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Doxorrubicina/farmacología , Células Hep G2 , Humanos , Células MCF-7 , Técnicas Analíticas Microfluídicas , Medicina de PrecisiónRESUMEN
Controllable manipulation of fluid flow is crucial for efficient particle separation, which is associated with plenty of biomedical and industrial applications. Microfluidic technologies have achieved promising progress in particle positioning depending on inertial force with or without the help of the Dean effect. Herein, we describe an inertial microfluidic system containing a spiral microchannel for various highly efficient particle separations. We demonstrated that Dean-like secondary flow can be regulated by geometric confinement in the microchannel. On the introduction of a library of micro-obstacles into the spiral microchannels, the resulting linear acceleration of secondary flow can be applied to remarkably enhance particle focusing in time and space. Further, multiple separating and sorting manipulations of particles including polymeric particles, circulating tumor cells, and blood cells, can be successfully accomplished in the dimension-confined spiral channels in a sheathless, high-throughput (typically 3 ml min-1), long-term (at least 4 h), and highly-efficient (up to 99.8% focusing) manner. The methodological achievement pointing to ease-of-use, effective, and high-throughput particle manipulations is useful for both laboratory and commercial developments of microfluidic systems in life and material sciences.
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Separación Celular/instrumentación , Técnicas Analíticas Microfluídicas/instrumentación , Animales , Células Sanguíneas/citología , Diseño de Equipo , Células HeLa , Humanos , Células K562 , Células MCF-7 , Ratones , Células Neoplásicas Circulantes/química , Tamaño de la PartículaRESUMEN
An Au nanoparticles/poly(caffeic acid) (AuNPs/PCA) composite modified glassy carbon (GC) electrode was prepared by successively potentiostatic technique in pH 7.4 phosphate buffer solution containing 0.02mM caffeic acid and 1.0mM HAuCl4. Electrochemical characterization of the AuNPs/PCA-GC electrode was investigated by electrochemical impedance spectroscopy and cyclic voltammetry. The electrochemical behavior of acetaminophen (AP) at the AuNPs/PCA-GC electrode was also studied by cyclic voltammetry. Compared with bare GC and poly(caffeic acid) modified GC electrode, the AuNPs/PCA-GC electrode was exhibited excellent electrocatalytic activity toward the oxidation of AP. The plot of catalytic current versus AP concentration showed two linear segments in the concentration ranges 0.2-20µM and 50-1000µM. The detection limit of 14 nM was obtained by using the first range of the calibration plot. The AuNPs/PCA-GC electrode has been successfully applied and validated by analyzing AP in blood, urine and pharmaceutical samples.
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Acetaminofén/análisis , Ácidos Cafeicos/química , Carbono/química , Técnicas de Química Analítica/instrumentación , Electrodos , Oro/química , Nanopartículas del Metal/química , Análisis Químico de la Sangre , Límite de Detección , Preparaciones Farmacéuticas/química , Orina/químicaRESUMEN
Engineering the liver in vitro is promising to provide functional replacement for patients with liver failure, or tissue models for drug metabolism and toxicity analysis. In this study, we describe a microfluidics-based biomimetic approach for the fabrication of an in vitro 3D liver lobule-like microtissue composed of a radially patterned hepatic cord-like network and an intrinsic hepatic sinusoid-like network. The hepatic enzyme assay showed that the 3D biomimetic microtissue maintained high basal CYP-1A1/2 and UGT activities, responded dynamically to enzyme induction/inhibition, and preserved great hepatic capacity of drug metabolism. Using the established biomimetic microtissue, the potential adverse drug reactions that induced liver injury were successfully analyzed via drug-drug interactions of clinical pharmaceuticals. The results showed that predosed pharmaceuticals which agitated CYP-1A1/2 and/or UGT activities would alter the toxic effect of the subsequently administrated drug. All the results validated the utility of the established biomimetic microtissue in toxicological studies in vitro. Also, we anticipate the microfluidics-based bioengineering strategy would benefit liver tissue engineering and liver physiology/pathophysiology studies, as well as in vitro assessment of drug-induced hepatotoxicity.
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Enfermedad Hepática Inducida por Sustancias y Drogas , Evaluación Preclínica de Medicamentos/métodos , Técnicas Analíticas Microfluídicas , Acetaminofén/efectos adversos , Acetaminofén/metabolismo , Acetaminofén/farmacología , Materiales Biomiméticos/química , Materiales Biomiméticos/metabolismo , Células Hep G2 , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Humanos , Isoniazida/efectos adversos , Isoniazida/metabolismo , Isoniazida/farmacología , Técnicas Analíticas Microfluídicas/instrumentación , Rifampin/efectos adversos , Rifampin/metabolismo , Rifampin/farmacología , Ingeniería de TejidosRESUMEN
Large-scale single-cell arrays are urgently required for current high-throughput screening of cell function and heterogeneity. However, the rapid and convenient generation of large-scale single-cell array in a multiplex and universal manner is not yet well established. In this paper, we report a simple and reliable method for the generation of a single-cell array by combining pneumatic microvalve arrays (PµVAs) and hydrodynamic single-cell trapping sites in a single microfluidic device. The PµVAs, which can be precisely controlled by actuated pressures, were designed to guide multiple types of cells being trapped in the corresponding single-cell trapping sites located in the fluidic channel. According to the theoretical demonstration and computational simulation, we successfully realized a multiplex single-cell array with three different types of cells by a step-by-step protocol. Furthermore, the analysis of cellular esterase heterogeneity of the three types of cells was concurrently implemented in the device as a proof-of-concept experiment. All the results demonstrated that the method developed in the current study could be applied for the generation of large-scale single-cell array with multiple cell types, which would be also promising and helpful for single-cell-based high-throughput drug test, multipurpose immunosensor and clinical diagnosis.
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Técnicas Biosensibles , Esterasas/aislamiento & purificación , Técnicas Analíticas Microfluídicas , Análisis de la Célula Individual , Línea Celular , Esterasas/química , Humanos , Hidrodinámica , PresiónRESUMEN
Cell sorting by filtration techniques offers a label-free approach for cell separation on the basis of size and deformability. However, filtration is always limited by the unpredictable variation of the filter hydrodynamic resistance due to cell accumulation and clogging in the microstructures. In this study, we present a new integrated microfluidic device for cell separation based on the cell size and deformability by combining the microstructure-constricted filtration and pneumatic microvalves. Using this device, the cell populations sorted by the microstructures can be easily released in real time for subsequent analysis. Moreover, the periodical sort and release of cells greatly avoided cell accumulation and clogging and improved the selectivity. Separation of cancer cells (MCF-7, MDA-MB-231 and MDA231-LM2) with different deformability showed that the mixture of the less flexible cells (MCF-7) and the flexible cells (MDA-MB-231 and MDA231-LM2) can be well separated with more than 75% purity. Moreover, the device can be used to separate cancer cells from the blood samples with more than 90% cell recovery and more than 80% purity. Compared with the current filtration methods, the device provides a new approach for cancer cell separation with high collection recovery and purity, and also, possesses practical potential to be applied as a sample preparation platform for fundamental studies and clinical applications.
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Separación Celular/métodos , Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas/instrumentación , Neoplasias/metabolismo , Neoplasias/patología , Línea Celular Tumoral , Separación Celular/instrumentación , Tamaño de la Célula , Supervivencia Celular , Diseño de Equipo , Filtración , Humanos , Hidrodinámica , Procesamiento de Imagen Asistido por Computador , Células MCF-7 , Técnicas Analíticas Microfluídicas/métodosRESUMEN
The development and application of miniaturized platforms with the capability for microscale and dynamic control of biomimetic and high-throughput three-dimensional (3D) culture plays a crucial role in biological research. In this study, pneumatic microstructure-based microfluidics was used to systematically demonstrate 3D tumor culture under various culture conditions. We also demonstrated the reusability of the fabrication-optimized pneumatic device for high-throughput cell manipulation and 3D tumor culture. This microfluidic system provides remarkably long-term (over 1 month) and cyclic stability. Furthermore, temporal and high-throughput monitoring of tumor response to evaluate the therapeutic efficacy of different chemotherapies, was achieved based on the robust culture. This advancement in microfluidics has potential applications in the fields of tissue engineering, tumor biology, and clinical medicine; it also provides new insight into the construction of high-performance and recyclable microplatforms for cancer research.
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Técnicas de Cultivo de Célula/instrumentación , Ensayos de Selección de Medicamentos Antitumorales/instrumentación , Técnicas Analíticas Microfluídicas/instrumentación , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Diseño de Equipo , Equipo Reutilizado , Ensayos Analíticos de Alto Rendimiento/instrumentación , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Reciclaje , Transducción de SeñalRESUMEN
The presence and quantity of rare cells in the bloodstream of cancer patients provide a potentially accessible source for the early detection of invasive cancer and for monitoring the treatment of advanced diseases. The separation of rare cells from peripheral blood, as a "virtual and real-time liquid biopsy", is expected to replace conventional tissue biopsies of metastatic tumors for therapy guidance. However, technical obstacles, similar to looking for a needle in a haystack, have hindered the broad clinical utility of this method. In this study, we developed a multistage microfluidic device for continuous label-free separation and enrichment of rare cells from blood samples based on cell size and deformability. We successfully separated tumor cells (MCF-7 and HeLa cells) and leukemic (K562) cells spiked in diluted whole blood using a unique complementary combination of inertial microfluidics and steric hindrance in a microfluidic system. The processing parameters of the inertial focusing and steric hindrance regions were optimized to achieve high-throughput and high-efficiency separation, significant advantages compared with existing rare cell isolation technologies. The results from experiments with rare cells spiked in 1% hematocrit blood indicated >90% cell recovery at a throughput of 2.24 × 10(7) cells min(-1). The enrichment of rare cells was >2.02 × 10(5)-fold. Thus, this microfluidic system driven by purely hydrodynamic forces has practical potential to be applied either alone or as a sample preparation platform for fundamental studies and clinical applications.