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BACKGROUND: Single-cell droplet microfluidics is an important platform for high-throughput analyses and screening because it provides an independent and compartmentalized microenvironment for reaction or cultivation by coencapsulating individual cells with various molecules in monodisperse microdroplets. In combination with microbial biosensors, this technology becomes a potent tool for the screening of mutant strains. In this study, we demonstrated that a genetically engineered yeast strain that can fluorescently sense agonist ligands via the heterologous expression of a human G-protein-coupled receptor (GPCR) and concurrently secrete candidate peptides is highly compatible with single-cell droplet microfluidic technology for the high-throughput screening of new agonistically active peptides. RESULTS: The water-in-oil microdroplets were generated using a flow-focusing microfluidic chip to encapsulate engineered yeast cells coexpressing a human GPCR [i.e., angiotensin II receptor type 1 (AGTR1)] and a secretory agonistic peptide [i.e., angiotensin II (Ang II)]. The single yeast cells cultured in the droplets were then observed under a microscope and analyzed using image processing incorporating machine learning techniques. The AGTR1-mediated signal transduction elicited by the self-secreted Ang II peptide was successfully detected via the expression of a fluorescent reporter in single-cell yeast droplet cultures. The system could also distinguish Ang II analog peptides with different agonistic activities. Notably, we further demonstrated that the microenvironment of the single-cell droplet culture enabled the detection of rarely existing positive (Ang II-secreting) yeast cells in the model mixed cell library, whereas the conventional batch-culture environment using a shake flask failed to do so. Thus, our approach provided compartmentalized microculture environments, which can prevent the diffusion, dilution, and cross-contamination of peptides secreted from individual single yeast cells for the easy identification of GPCR agonists. CONCLUSIONS: We established a droplet-based microfluidic platform that integrated an engineered yeast biosensor strain that concurrently expressed GPCR and self-secreted the agonistic peptides. This offers individually isolated microenvironments that allow the culture of single yeast cells secreting these peptides and gaging their signaling activities, for the high-throughput screening of agonistic peptides. Our platform base on yeast GPCR biosensors and droplet microfluidics will be widely applicable to metabolic engineering, environmental engineering, and drug discovery.
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Microfluídica , Saccharomyces cerevisiae , Humanos , Microfluídica/métodos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Péptidos/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Ensayos Analíticos de Alto RendimientoRESUMEN
Cell surface properties of microorganisms provide abundant information for their physiological status and fate choice. However, current methods for analyzing cell surface properties require labeling or fixation, which can alter the cell activity. This study establishes a label-free, rapid, noninvasive, and quantitative analysis of cell surface properties, including the presence and the dimension of epistructure, down to the single-cell level and at the nanometer scale. Simultaneously, electrorotation provides dielectric properties of intracellular contents. With the combined information, the growth phase of microalgae cells can be identified. The measurement is based on electrorotation of single cells, and an electrorotation model accounting for the surface properties is developed to properly interpret experimental data. The epistructure length measured by electrorotation is validated by scanning electron microscopy. The measurement accuracy is satisfactory in particular in the case of microscale epistructures in the exponential phase and nanoscale epistructures in the stationary phase. However, the measurement accuracy for nanoscale epistructures on cells in the exponential phase is offset by the effect of a thick double layer. Lastly, a diversity in epistructure length distinguishes exponential phase from stationary phase.
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Membrana CelularRESUMEN
The load on drive elements under extreme pressure conditions is significantly larger than that used in machine tools. When operating under a heavy load for a long period, large deformation and severe wear between the ball and the track are more likely to occur. To reduce wear, the most fundamental solution is to improve the surface properties of the material. Moreover, heat treatment is the most effective method to improve the surface properties of materials, thereby achieving wear resistance and low friction. It is necessary to develop a new heat treatment technology for wear resistance in extreme pressure conditions. Therefore, this study conducted experiments using a reciprocating friction tester. The responses of electrical contact resistance and the friction coefficient were measured synchronously to investigate wear resistance and low friction of the alloy steels after the induction heat treatment. Then, the results were compared and verified with low-carbon alloy steel after the traditional carburizing heat treatment. The experimental results show that the application of new induction heat treatment technology can not only improve the performance of drive components, but also save time and energy, and streamline the production process of the drive components. Therefore, the results of these wear analyses confirm that the induction heat treatment mode can replace the traditional carburizing heat treatment mode for drive elements.
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Photosynthetic microalgae not only perform fixation of carbon dioxide but also produce valuable byproducts such as lipids and pigments. However, due to the lack of effective tools for rapid and noninvasive analysis of microalgal cellular contents, the efficiency of strain screening and culture optimizing is usually quite low. This study applied single-cell electrorotation on Scenedesmus abundans to assess cellular dielectric properties during lipid accumulation and to promptly quantify total cellular contents. The experimental electrorotation spectra were fitted with the double-shell ellipsoidal model, which considered varying cell wall thickness, to obtain the dielectric properties of cellular compartments. When the amount of total lipids increased from 15.3 wt% to 33.8 wt%, the conductivity and relative permittivity of the inner core (composed of the cytoplasm, lipid droplets, and nucleus) decreased by 21.7% and 22.5%, respectively. These dielectric properties were further used to estimate the total cellular lipid contents by the general mixing formula, and the estimated values agreed with those obtained by weighing dry biomass and extracted lipids with an error as low as 0.22 wt%. Additionally, the conductivity and relative permittivity of cell wall increased during nitrogen-starvation conditions, indicating the thickening of cell wall, which was validated by the transmission electron microscopy.
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This study aims to investigate the efficacy of lipid induction in Scenedesmus abundans by adding the effluent from dark fermentation of biohydrogen production. Four sets of experiments were conducted: control (sufficient nitrogen), nitrogen depletion, low concentration (0.3×) effluent addition, and high concentration (0.5×) effluent addition. The addition of low concentration effluent produced the highest biomass and lipid yields of 2.831 g/L and 1.238 g/L, corresponding to a lipid abundance of 43.72 wt%. Furthermore, S. abundans had high removal efficiencies for volatile fatty acids in the effluent (formic acid 100%, acetic acid 100%, propionic acid 98%, lactic acid 84%, and butyric acid 68%), and this is the first study demonstrating the ability of S. abundans in using formic acid and lactic acid to produce biomass and lipids. These results show that S. abundans have great abilities in simultaneous reducing organic acids in the effluent and producing valuable metabolites.
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Biocombustibles , Ácidos Grasos Volátiles/metabolismo , Hidrógeno/metabolismo , Metabolismo de los Lípidos , Scenedesmus/crecimiento & desarrollo , AnaerobiosisRESUMEN
A key challenge for bioprocess engineering is the identification of the optimum process conditions for the production of biochemical and biopharmaceutical compounds using prokaryotic as well as eukaryotic cell factories. Shake flasks and bench-scale bioreactor systems are still the golden standard in the early stage of bioprocess development, though they are known to be expensive, time-consuming, and labor-intensive as well as lacking the throughput for efficient production optimizations. To bridge the technological gap between bioprocess optimization and upscaling, we have developed a microfluidic bioreactor array to reduce time and costs, and to increase throughput compared with traditional lab-scale culture strategies. We present a multifunctional microfluidic device containing 12 individual bioreactors (Vt = 15 µl) in a 26 mm × 76 mm area with in-line biosensing of dissolved oxygen and biomass concentration. Following initial device characterization, the bioreactor lab-on-a-chip was used in a proof-of-principle study to identify the most productive cell line for lactic acid production out of two engineered yeast strains, evaluating whether it could reduce the time needed for collecting meaningful data compared with shake flasks cultures. Results of the study showed significant difference in the strains' productivity within 3 hr of operation exhibiting a 4- to 6-fold higher lactic acid production, thus pointing at the potential of microfluidic technology as effective screening tool for fast and parallelizable industrial bioprocess development.
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Reactores Biológicos , Ácido Láctico/metabolismo , Saccharomyces cerevisiae/metabolismo , Diseño de Equipo , Microbiología Industrial/instrumentación , Dispositivos Laboratorio en un Chip , Saccharomyces cerevisiae/citologíaRESUMEN
This review presents a critical assessment of emerging microfluidic technologies for the application on biological productions of biofuels and other chemicals from microalgae. Comparisons of cell culture designs for the screening of microalgae strains and growth conditions are provided with three categories: mechanical traps, droplets, or microchambers. Emerging technologies for the in situ characterization of microalgae features and metabolites are also presented and evaluated. Biomass and secondary metabolite productivities obtained at microscale are compared with the values obtained at bulk scale to assess the feasibility of optimizing large-scale operations using microfluidic platforms. The recent studies in microsystems for microalgae pretreatment, fractionation and extraction of metabolites are also reviewed. Finally, comments toward future developments (high-pressure/-temperature process; solvent-resistant devices; omics analysis, including genome/epigenome, proteome, and metabolome; biofilm reactors) of microfluidic techniques for microalgae applications are provided.
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Microfluidic systems based on fluorescence detection have been developed and applied for many biological and chemical applications. Because of the tiny amount of sample in the system; the induced fluorescence can be weak. Therefore, most microfluidic systems deploy multiple optical components or sophisticated equipment to enhance the efficiency of fluorescence detection. However, these strategies encounter common issues of complex manufacturing processes and high costs. In this study; a miniature, cylindrical and hybrid lens made of polydimethylsiloxane (PDMS) to improve the fluorescence detection in microfluidic systems is proposed. The hybrid lens integrates a laser focusing lens and a fluorescence collecting lens to achieve dual functions and simplify optical setup. Moreover, PDMS has advantages of low-cost and straightforward fabrication compared with conventional optical components. The performance of the proposed lens is first examined with two fluorescent dyes and the results show that the lens provides satisfactory enhancement for fluorescence detection of Rhodamine 6G and Nile Red. The overall increments in collected fluorescence signal and detection sensitivity are more than 220% of those without lens, and the detection limits of Rhodamine 6G and Nile red are lowered to 0.01 µg/mL and 0.05 µg/mL, respectively. The hybrid lens is further applied to the detection of Nile red-labeled Chlorella vulgaris cells and it increases both signal intensity and detection sensitivity by more than 520%. The proposed hybrid lens also dramatically reduces the variation in detected signal caused by the deviation in incident angle of excitation light.
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Microalgae have emerged as one of the most promising feedstocks for biofuels and bio-based chemical production. However, due to the lack of effective tools enabling rapid and high-throughput analysis of the content of microalgae biomass, the efficiency of screening and identification of microalgae with desired functional components from the natural environment is usually quite low. Moreover, the real-time monitoring of the production of target components from microalgae is also difficult. Recently, research efforts focusing on overcoming this limitation have started. In this review, the recent development of high-throughput methods for analyzing microalgae cellular contents is summarized. The future prospects and impacts of these detection methods in microalgae-related processing and industries are also addressed.
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Biocombustibles/análisis , Ensayos Analíticos de Alto Rendimiento/métodos , Microalgas/metabolismo , Biomasa , Biotecnología/métodos , Carbohidratos/análisis , Ensayos Analíticos de Alto Rendimiento/instrumentación , Lípidos/análisis , Proteínas/análisisRESUMEN
Chlorella vulgaris ESP-31 containing 22.7% lipid was harvested by coagulation (using chitosan and polyaluminium chloride (PACl) as the coagulants) and centrifugation. The harvested ESP-31 was directly employed as the oil source for biodiesel production via transesterification catalyzed by immobilized Burkholderia lipase and by a synthesized solid catalyst (SrO/SiO2). Both enzymatic and chemical transesterification were significantly inhibited in the presence of PACl, while the immobilized lipase worked well with wet chitosan-coagulated ESP-31, giving a high biodiesel conversion of 97.6% w/w oil, which is at a level comparable to that of biodiesel conversion from centrifugation-harvested microalgae (97.1% w/w oil). The immobilized lipase can be repeatedly used for three cycles without significant loss of its activity. The solid catalyst SrO/SiO2 worked well with water-removed centrifuged ESP-31 with a biodiesel conversion of 80% w/w oil, but the conversion became lower (55.7-61.4% w/w oil) when using water-removed chitosan-coagulated ESP-31 as the oil source.
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Biocombustibles , Biotecnología/métodos , Chlorella vulgaris/crecimiento & desarrollo , Microalgas/crecimiento & desarrollo , Hidróxido de Aluminio/farmacología , Biomasa , Burkholderia/efectos de los fármacos , Burkholderia/enzimología , Catálisis/efectos de los fármacos , Centrifugación , Quitosano/farmacología , Chlorella vulgaris/efectos de los fármacos , Chlorella vulgaris/metabolismo , Enzimas Inmovilizadas/metabolismo , Esterificación/efectos de los fármacos , Floculación/efectos de los fármacos , Concentración de Iones de Hidrógeno/efectos de los fármacos , Lipasa/metabolismo , Lípidos/química , Microalgas/efectos de los fármacos , Microalgas/metabolismo , Tamaño de la Partícula , Dióxido de Silicio/farmacología , Electricidad Estática , Estroncio/farmacología , Factores de TiempoRESUMEN
A membraneless microfluidic microbial fuel cell (µMFC) for rapid detection of microorganism electroactivity is demonstrated in this study. Owing to the merit of laminar flow, the proposed µMFC has well-separated anode and cathode without applying proton exchange membrane. The highest open circuit voltages (OCVs) produced by different anodal solutions: fresh medium, inactivated and untreated microflora, were 102, 131, and 246 mV, respectively. These results show that the membraneless µMFC is capable of identifying the electric potential resulting from the imbalanced compositions between two streams (29 mV) and from the electrochemical activity of microflora (115 mV). When samples obtained along a batch cycle of H-type MFC were tested, the membraneless µMFC produced similar OCVs with those from the H-type MFC. In conclusion, the proposed µMFC has comparable abilities in detecting electroactivity with the conventional H-type MFC; moreover, it can distinguish the source of collected electricity.
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Fuentes de Energía Bioeléctrica/microbiología , Fenómenos Electrofisiológicos/fisiología , Técnicas Analíticas Microfluídicas/métodos , Electrodos , Taiwán , Factores de TiempoRESUMEN
A rapid and noninvasive quantification method for cellular lipids in Chlorella vulgaris is demonstrated in this study. This method applied near-infrared Raman spectroscopy to monitor the change of signal intensities at 1440 cm(-1) and 2845-3107 cm(-1) along the nitrogen depletion period, and calibration curves relating signal intensity and cellular lipid abundance were established. The calibration curves show that signal intensity at 2845-3107 cm(-1) and cellular lipid abundance were highly correlated. When the calibration curve was applied on the lipid quantification of two unknown samples, the differences between lipid abundances estimated by the calibration curve and measured by gas chromatography were less than 2 wt %. Carotenoids produced a strong and broad peak near 1440 cm(-1), and it weakened the correlation between signal intensity and lipid abundance. The consistency of detection and effects of cellular contents and water on the Raman spectrogram of Chlorella vulgaris were also addressed. The sample pretreatment only involved centrifugation, and the time required for lipid quantification was shortened to less than 1.5 h. The rapid detection has great potential in high-throughput screening of microalgae and also provides valuable information for monitoring the quality of microalgae culture and determining parameters for the mass production of biodiesel from microalgae.
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Chlorella vulgaris/metabolismo , Lípidos/análisis , Espectroscopía Infrarroja Corta , Celulosa/análisis , Aceite de Oliva , Pectinas/análisis , Aceites de Plantas/análisis , beta Caroteno/análisisRESUMEN
Reducing the fluorescence background in microfluidic assays is important in obtaining accurate outcomes and enhancing the quality of detections. This study demonstrates an integrated process including cell labelling, fluorescence background reduction, and biomolecule detection using liquid-liquid extraction in a microfluidic droplet system. The cellular lipids in Chlorella vulgaris and NIH/3T3 cells were labelled with a hydrophobic dye, Nile red, to investigate the performance of the proposed method. The fluorescence background of the lipid detection can be reduced by 85% and the removal efficiency increased with the volume of continuous phase surrounding a droplet. The removal rate of the fluorescence background increased as the surface area to volume ratio of a droplet increased. Before Nile red was removed from the droplet, the signal to noise ratio was as low as 1.30 and it was difficult to distinguish cells from the background. Removing Nile red increased the signal to noise ratio to 22 and 34 for Chlorella vulgaris and NIH/3T3, respectively, and these were 17 fold and 10 fold of the values before extraction. The proposed method successfully demonstrates the enhancement of fluorescence detection of cellular lipids and has great potential in improving other fluorescence-based detections in microfluidic systems.
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Fluorescencia , Lípidos/análisis , Extracción Líquido-Líquido/métodos , Técnicas Analíticas Microfluídicas/métodos , Animales , Células Cultivadas , Chlorella vulgaris/química , Extracción Líquido-Líquido/instrumentación , Ratones , Técnicas Analíticas Microfluídicas/instrumentación , Células 3T3 NIH , Oxazinas/química , Tamaño de la PartículaRESUMEN
An alternating current was used to generate an electric field to enhance the fluorescent labeling of microalgae cellular lipids with Nile red and LipidTOX. The decay of the fluorescence intensity of Chlorella vulgaris cells in 0 V/cm was more than 50% after 10 min, and the intensity variation was as high as 7% in 20s. At 2000 V/cm, the decay rate decreased to 1.22% per minute and the intensity fluctuation was less than 1% for LipidTOX-labeled cells. For Spirulina sp. cells at 0 V/cm, the fluorescence intensity increased by 10% after 10 min, whereas at 2000 V/cm, labeling was more rapid and fluorescence intensity doubled. These results show that applying an electric field can improve the quality of fluorescence detection by alleviating decay and fluctuation or by enhancing signal intensity.
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Electricidad , Microalgas/metabolismo , FluorescenciaRESUMEN
This review presents the development of micro-sized microbial fuel cells (including mL-scale and µL-scale setups), with summarization of their advantageous characteristics, fabrication methods, performances, potential applications and possible future directions. The performance of microbial fuel cells (MFCs) is affected by issues such as mass transport, reaction kinetics and ohmic resistance. These factors are manipulated in micro-sized MFCs using specially allocated electrodes constructed with specified materials having physically or chemically modified surfaces. Both two-chamber and air-breathing cathodes are promising configurations for mL-scale MFCs. However, most of the existing µL-scale MFCs generate significantly lower volumetric power density compared with their mL-counterparts because of the high internal resistance. Although µL-scale MFCs have not yet to provide sufficient power for operating conventional equipment, they show great potential in rapid screening of electrochemically microbes and electrode performance. Additional possible applications and future directions are also provided for the development of micro-sized MFCs.
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Bacterias/metabolismo , Fuentes de Energía Bioeléctrica/microbiología , Microquímica/instrumentación , Aire , Bacterias/genética , Impedancia Eléctrica , Electrodos/microbiología , Predicción , MiniaturizaciónRESUMEN
Genetic modification of cells is a critical step involved in many cell therapy and gene therapy protocols. In these applications, cell samples of large volume (10(8)-10(9)cells) are often processed for transfection. This poses new challenges for current transfection methods and practices. Here we present a novel flow-through electroporation method for delivery of genes into cells at high flow rates (up to approximately 20 mL/min) based on disposable microfluidic chips, a syringe pump, and a low-cost direct current (DC) power supply that provides a constant voltage. By eliminating pulse generators used in conventional electroporation, we dramatically lowered the cost of the apparatus and improved the stability and consistency of the electroporation field for long-time operation. We tested the delivery of pEFGP-C1 plasmids encoding enhanced green fluorescent protein into Chinese hamster ovary (CHO-K1) cells in the devices of various dimensions and geometries. Cells were mixed with plasmids and then flowed through a fluidic channel continuously while a constant voltage was established across the device. Together with the applied voltage, the geometry and dimensions of the fluidic channel determined the electrical parameters of the electroporation. With the optimal design, approximately 75% of the viable CHO cells were transfected after the procedure. We also generalize the guidelines for scaling up these flow-through electroporation devices. We envision that this technique will serve as a generic and low-cost tool for a variety of clinical applications requiring large volume of transfected cells.
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Electroporación/instrumentación , Electroporación/métodos , Animales , Células CHO , Células , Cricetinae , Cricetulus , Electricidad , Proteínas Fluorescentes Verdes , Microfluídica/métodos , Fenómenos Físicos , Plásmidos , Transfección/instrumentaciónRESUMEN
To elucidate the Tau gain-of-toxicity functional mechanism and to search for potential treatments, we overexpressed human Tau variants (hTau) in the dorsal mesothorax (notum) of Drosophila. Overexpression of Tau variants caused loss of notal bristles, and the phenotype was used for evaluating toxicity of ectopic Tau. The bristle loss phenotype was found to be highly associated with the toxicity of hyperphosphoryled Tau in flies. We have shown that the bristle loss phenotype can be rescued either by reducing Glycogen synthase kinase 3beta (GSK3beta)/Shaggy (Sgg) activity or overexpressing Bbeta2 regulatory subunits of PP2A. Elevated expression of the Drosophila Bbeta2 homolog, Twins (Tws), also alleviated neuritic dystrophy of the dorsal arborization (da) neuron caused by Tau aggregation. Additionally, lowering endogenous Tau dosage was beneficial as it ameliorated the bristle loss phenotype. Finally, the bristle loss phenotype was used to evaluate the efficacy of potential therapeutic compounds. The GSK3beta inhibitor, alsterpaullone, was found to suppress toxicity of Tau in a concentration-dependent manner. The notum of Drosophila, thus, provides a new tool and insights into Tau-induced toxicity. It could also potentially assist in screening new drugs for possible therapeutic intervention.
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Animales Modificados Genéticamente , Drosophila melanogaster/genética , Glucógeno Sintasa Quinasa 3/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/aislamiento & purificación , Tauopatías/metabolismo , Proteínas tau/metabolismo , Animales , Benzazepinas/farmacología , Modelos Animales de Enfermedad , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Glucógeno Sintasa Quinasa 3/metabolismo , Humanos , Indoles/farmacología , Ovillos Neurofibrilares/metabolismo , Fosfoproteínas Fosfatasas/metabolismo , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Proteínas tau/genéticaRESUMEN
Microchip CE of proteins labeled either off- or on-chip with the "chameleon" CE dye 503 using poly(methyl methacrylate) microchips is presented. A simple dynamic coating using the cationic surfactant CTAB prevented nonspecific adsorption of protein and dye to the channel walls. The labeling reactions for both off- and on-chip labeling proceeded at room temperature without requiring heating steps. In off-chip labeling, a 9 ng/mL concentration detection limit for BSA, corresponding to a approximately 7 fg (100 zmol) mass detection limit, was obtained. In on-chip tagging, the free dye and protein were placed in different reservoirs of the microchip, and an extra incubation step was not needed. A 1 microg/mL concentration detection limit for BSA, corresponding to a approximately 700 fg (10 amol) mass detection limit, was obtained from this protocol. The earlier elution time of the BSA peak in on-chip labeling resulted from fewer total labels on each protein molecule. Our on-chip labeling method is an important part of automation in miniaturized devices.
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Colorantes/química , Electroforesis Capilar/métodos , Dispositivos Laboratorio en un Chip , Polímeros/química , Adsorción , Límite de Detección , Microscopía Fluorescente , Miniaturización , Albúmina Sérica Bovina/análisisRESUMEN
Detection and accurate quantitation of biomarkers such as alpha-fetoprotein (AFP) can be a key aspect of early stage cancer diagnosis. Microfluidic devices provide attractive analysis capabilities, including low sample and reagent consumption, as well as short assay times. However, to date microfluidic analyzers have relied almost exclusively on calibration curves for sample quantitation, which can be problematic for complex mixtures such as human serum. We have fabricated integrated polymer microfluidic systems that can quantitatively determine fluorescently labeled AFP in human serum using either the method of standard addition or a calibration curve. Our microdevices couple an immunoaffinity purification step with rapid microchip electrophoresis separation in a laser-induced fluorescence detection system, all under automated voltage control in a miniaturized polymer microchip. In conjunction with laser-induced fluorescence detection, these systems can quantify AFP at approximately 1 ng/mL levels in approximately 10 microL of human serum in a few tens of minutes. Our polymer microdevices have been applied in determining AFP in spiked serum samples. These integrated microsystems offer excellent potential for rapid, simple, and accurate biomarker quantitation in a point-of-care setting.
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Colorantes Fluorescentes/química , Técnicas Analíticas Microfluídicas/instrumentación , alfa-Fetoproteínas/análisis , Calibración , Cromatografía de Afinidad , Detección Precoz del Cáncer , Electroforesis por Microchip , Humanos , Técnicas Analíticas Microfluídicas/métodos , Técnicas Analíticas Microfluídicas/normas , Polímeros/química , Factores de TiempoRESUMEN
Microfluidic arrays of living cells have raised a lot of interests recently due to their potential for high throughput screening of cell-based assays. This report presents a microfluidic cell array with individually addressable chambers controlled by pneumatic valves for cell culture and treatment. There are two modes for the cell array to be operated. In the first mode, different groups of cells are directed into designated chambers for culturing and observation. We demonstrate the delivery and culture of enhanced green fluorescent protein (EGFP) expressing and nonfluorescent Chinese hamster ovary (CHO) cells into specific chambers in the array. In the second mode, the chambers are first seeded with the same cell type and different reagents are delivered to specific chambers for cell treatment. We treat cells in designated chambers with Calcein AM and CellTrace calcein red-orange AM to demonstrate the principle. We envision that this microfluidic cell array technology will pave the way to automated high-throughput screening of biomolecules and drugs based on observing cellular phenotypes and responses.