The Experts below are selected from a list of 252 Experts worldwide ranked by ideXlab platform
Michael T. Bowser - One of the best experts on this subject based on the ideXlab platform.
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reduced surface adsorption in 3d printed acrylonitrile butadiene styrene micro Free Flow Electrophoresis devices
Electrophoresis, 2020Co-Authors: Sarah K. Anciaux, Michael T. BowserAbstract:We have 3D printed and fabricated micro Free-Flow Electrophoresis (µFFE) devices in acrylonitrile butadiene styrene (ABS) that exhibit minimal surface adsorption without requiring additional surface coatings or specialized buffer additives. 2D, nano LC-micro Free Flow Electrophoresis (2D nLC × µFFE) separations were used to assess both spatial and temporal broadening as peaks eluted through the separation channel. Minimal broadening due to wall adsorption was observed in either the spatial or temporal dimensions during separations of rhodamine 110, rhodamine 123, and fluorescein. Surface adsorption was observed in separations of Chromeo P503 labeled myoglobin and cytochrome c but was significantly reduced compared to previously reported glass devices. Peak widths of 20 min. A 2D nLC × µFFE separation of a Chromeo P503 labeled tryptic digest of BSA was performed to demonstrate the high peak capacity possible due to the low surface adsorption in the 3D printed ABS devices, even in the absence of surface coatings or buffer additives.
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Reduced surface adsorption in 3D printed acrylonitrile butadiene styrene micro Free‐Flow Electrophoresis devices
Electrophoresis, 2019Co-Authors: Sarah K. Anciaux, Michael T. BowserAbstract:We have 3D printed and fabricated micro Free-Flow Electrophoresis (µFFE) devices in acrylonitrile butadiene styrene (ABS) that exhibit minimal surface adsorption without requiring additional surface coatings or specialized buffer additives. 2D, nano LC-micro Free Flow Electrophoresis (2D nLC × µFFE) separations were used to assess both spatial and temporal broadening as peaks eluted through the separation channel. Minimal broadening due to wall adsorption was observed in either the spatial or temporal dimensions during separations of rhodamine 110, rhodamine 123, and fluorescein. Surface adsorption was observed in separations of Chromeo P503 labeled myoglobin and cytochrome c but was significantly reduced compared to previously reported glass devices. Peak widths of 20 min. A 2D nLC × µFFE separation of a Chromeo P503 labeled tryptic digest of BSA was performed to demonstrate the high peak capacity possible due to the low surface adsorption in the 3D printed ABS devices, even in the absence of surface coatings or buffer additives.
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Micro Free Flow Electrophoresis
Lab on a chip, 2017Co-Authors: Alex C. Johnson, Michael T. BowserAbstract:Micro Free-Flow Electrophoresis (μFFE) is a continuous separation technique in which analytes are streamed through a perpendicularly applied electric field in a planar separation channel. Analyte streams are deflected laterally based on their electrophoretic mobilities as they Flow through the separation channel. A number of μFFE separation modes have been demonstrated, including Free zone (FZ), micellar electrokinetic chromatography (MEKC), isoelectric focusing (IEF) and isotachophoresis (ITP). Approximately 60 articles have been published since the first μFFE device was fabricated in 1994. We anticipate that recent advances in device design, detection, and fabrication, will allow μFFE to be applied to a much wider range of applications. Applications particularly well suited for μFFE analysis include continuous, real time monitoring and microscale purifications.
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3D Printed Micro Free-Flow Electrophoresis Device
Analytical Chemistry, 2016Co-Authors: Sarah K. Anciaux, Matthew Geiger, Michael T. BowserAbstract:The cost, time, and restrictions on creative flexibility associated with current fabrication methods present significant challenges in the development and application of microfluidic devices. Additive manufacturing, also referred to as three-dimensional (3D) printing, provides many advantages over existing methods. With 3D printing, devices can be made in a cost-effective manner with the ability to rapidly prototype new designs. We have fabricated a micro Free-Flow Electrophoresis (μFFE) device using a low-cost, consumer-grade 3D printer. Test prints were performed to determine the minimum feature sizes that could be reproducibly produced using 3D printing fabrication. Microfluidic ridges could be fabricated with dimensions as small as 20 μm high × 640 μm wide. Minimum valley dimensions were 30 μm wide × 130 μm wide. An acetone vapor bath was used to smooth acrylonitrile–butadiene–styrene (ABS) surfaces and facilitate bonding of fully enclosed channels. The surfaces of the 3D-printed features were profile...
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3D Printed Micro Free-Flow Electrophoresis Device
2016Co-Authors: Sarah K. Anciaux, Matthew Geiger, Michael T. BowserAbstract:The cost, time, and restrictions on creative flexibility associated with current fabrication methods present significant challenges in the development and application of microfluidic devices. Additive manufacturing, also referred to as three-dimensional (3D) printing, provides many advantages over existing methods. With 3D printing, devices can be made in a cost-effective manner with the ability to rapidly prototype new designs. We have fabricated a micro Free-Flow Electrophoresis (μFFE) device using a low-cost, consumer-grade 3D printer. Test prints were performed to determine the minimum feature sizes that could be reproducibly produced using 3D printing fabrication. Microfluidic ridges could be fabricated with dimensions as small as 20 μm high × 640 μm wide. Minimum valley dimensions were 30 μm wide × 130 μm wide. An acetone vapor bath was used to smooth acrylonitrile–butadiene–styrene (ABS) surfaces and facilitate bonding of fully enclosed channels. The surfaces of the 3D-printed features were profiled and compared to a similar device fabricated in a glass substrate. Stable stream profiles were obtained in a 3D-printed μFFE device. Separations of fluorescent dyes in the 3D-printed device and its glass counterpart were comparable. A μFFE separation of myoglobin and cytochrome c was also demonstrated on a 3D-printed device. Limits of detection for rhodamine 110 were determined to be 2 and 0.3 nM for the 3D-printed and glass devices, respectively
Detlev Belder - One of the best experts on this subject based on the ideXlab platform.
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Nonaqueous Micro Free-Flow Electrophoresis for Continuous Separation of Reaction Mixtures in Organic Media.
Analytical Chemistry, 2019Co-Authors: Benjamin M. Rudisch, Simon A. Pfeiffer, David Geissler, Elisabeth Speckmeier, Andrea A. Robitzki, Kirsten Zeitler, Detlev BelderAbstract:The continuous separation mechanism of micro Free-Flow Electrophoresis (μFFE) is a straightforward, suitable tool for microscale purification of reaction mixtures. However, aqueous separation buffers and organic reaction solvents limit the applicability of this promising combination. Herein, we have explored nonaqueous micro Free-Flow Electrophoresis for this purpose and present its suitability for a continuous workup of organic reactions performed in acetonitrile. After successful nonaqueous FFE separation of organic dyes, the approach was applied to continuously recover the photocatalyst [Ru(bpy)3]2+ from a homogeneous, acetonitrile-based reaction mixture. This approach opens up possibilities for further downstream processing of purified products and is also attractive for recycling of precious catalyst species.
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Nonaqueous Micro Free-Flow Electrophoresis for Continuous Separation of Reaction Mixtures in Organic Media
2019Co-Authors: Benjamin M. Rudisch, Simon A. Pfeiffer, David Geissler, Elisabeth Speckmeier, Andrea A. Robitzki, Kirsten Zeitler, Detlev BelderAbstract:The continuous separation mechanism of micro Free-Flow Electrophoresis (μFFE) is a straightforward, suitable tool for microscale purification of reaction mixtures. However, aqueous separation buffers and organic reaction solvents limit the applicability of this promising combination. Herein, we have explored nonaqueous micro Free-Flow Electrophoresis for this purpose and present its suitability for a continuous workup of organic reactions performed in acetonitrile. After successful nonaqueous FFE separation of organic dyes, the approach was applied to continuously recover the photocatalyst [Ru(bpy)3]2+ from a homogeneous, acetonitrile-based reaction mixture. This approach opens up possibilities for further downstream processing of purified products and is also attractive for recycling of precious catalyst species
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Chip-based Free-Flow Electrophoresis with integrated nanospray mass-spectrometry.
Angewandte Chemie (International ed. in English), 2015Co-Authors: Christian Benz, Michael Boomhoff, Johannes Appun, Christoph Schneider, Detlev BelderAbstract:Free-Flow Electrophoresis is an ideal tool for preparative separations in continuous microFlow. With the approach presented herein for coupling Free-Flow Electrophoresis and mass spectrometry it is now also possible to trace non-fluorescent compounds and identify them by means of mass spectrometry. The functionality of the method and its potential as an integrated separation unit for microFlow synthesis is demonstrated by application to a multicomponent [3+2]-cycloannulation.
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Towards an integrated device that utilizes adherent cells in a micro-Free-Flow Electrophoresis chip to achieve separation and biosensing
Analytical and Bioanalytical Chemistry, 2013Co-Authors: Stefan Jezierski, Christian Benz, Anke S. Klein, Michael Schaefer, Stefan Nagl, Detlev BelderAbstract:We immobilized adherent human embryonic kidney (HEK) cells—which are able to trace adenosine triphosphate (ATP) —inside a microfluidic Free-Flow Electrophoresis (μFFE) chip in order to develop an integrated device combining separation and biosensing capabilities. HEK 293 cells loaded with fluorescent calcium indicators were used as a model system to enable the spatially and temporally resolved detection of ATP. The local position of a 20 μM ATP stream was successfully visualized by these cells during Free-Flow Electrophoresis, demonstrating the on-line detection capability of this technique towards native, unlabeled compounds.
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label Free real time imaging in microchip Free Flow Electrophoresis applying high speed deep uv fluorescence scanning
Lab on a Chip, 2012Co-Authors: Stefan Köhler, Stefan Nagl, Stefanie Fritzsche, Detlev BelderAbstract:We report on label-Free monitoring of microfluidic Free-Flow Electrophoresis (μFFE) separations in real-time using a custom built high speed deep UV laser scanner. In combination with a novel layout realized in fused silica (FS) FFE chips the setup was successfully applied for continuous separations and detection of unlabeled analytes including native proteins by space-resolved intrinsic deep UV fluorescence scanning.
Sarah K. Anciaux - One of the best experts on this subject based on the ideXlab platform.
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reduced surface adsorption in 3d printed acrylonitrile butadiene styrene micro Free Flow Electrophoresis devices
Electrophoresis, 2020Co-Authors: Sarah K. Anciaux, Michael T. BowserAbstract:We have 3D printed and fabricated micro Free-Flow Electrophoresis (µFFE) devices in acrylonitrile butadiene styrene (ABS) that exhibit minimal surface adsorption without requiring additional surface coatings or specialized buffer additives. 2D, nano LC-micro Free Flow Electrophoresis (2D nLC × µFFE) separations were used to assess both spatial and temporal broadening as peaks eluted through the separation channel. Minimal broadening due to wall adsorption was observed in either the spatial or temporal dimensions during separations of rhodamine 110, rhodamine 123, and fluorescein. Surface adsorption was observed in separations of Chromeo P503 labeled myoglobin and cytochrome c but was significantly reduced compared to previously reported glass devices. Peak widths of 20 min. A 2D nLC × µFFE separation of a Chromeo P503 labeled tryptic digest of BSA was performed to demonstrate the high peak capacity possible due to the low surface adsorption in the 3D printed ABS devices, even in the absence of surface coatings or buffer additives.
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Reduced surface adsorption in 3D printed acrylonitrile butadiene styrene micro Free‐Flow Electrophoresis devices
Electrophoresis, 2019Co-Authors: Sarah K. Anciaux, Michael T. BowserAbstract:We have 3D printed and fabricated micro Free-Flow Electrophoresis (µFFE) devices in acrylonitrile butadiene styrene (ABS) that exhibit minimal surface adsorption without requiring additional surface coatings or specialized buffer additives. 2D, nano LC-micro Free Flow Electrophoresis (2D nLC × µFFE) separations were used to assess both spatial and temporal broadening as peaks eluted through the separation channel. Minimal broadening due to wall adsorption was observed in either the spatial or temporal dimensions during separations of rhodamine 110, rhodamine 123, and fluorescein. Surface adsorption was observed in separations of Chromeo P503 labeled myoglobin and cytochrome c but was significantly reduced compared to previously reported glass devices. Peak widths of 20 min. A 2D nLC × µFFE separation of a Chromeo P503 labeled tryptic digest of BSA was performed to demonstrate the high peak capacity possible due to the low surface adsorption in the 3D printed ABS devices, even in the absence of surface coatings or buffer additives.
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3D Printed Micro Free-Flow Electrophoresis Device
Analytical Chemistry, 2016Co-Authors: Sarah K. Anciaux, Matthew Geiger, Michael T. BowserAbstract:The cost, time, and restrictions on creative flexibility associated with current fabrication methods present significant challenges in the development and application of microfluidic devices. Additive manufacturing, also referred to as three-dimensional (3D) printing, provides many advantages over existing methods. With 3D printing, devices can be made in a cost-effective manner with the ability to rapidly prototype new designs. We have fabricated a micro Free-Flow Electrophoresis (μFFE) device using a low-cost, consumer-grade 3D printer. Test prints were performed to determine the minimum feature sizes that could be reproducibly produced using 3D printing fabrication. Microfluidic ridges could be fabricated with dimensions as small as 20 μm high × 640 μm wide. Minimum valley dimensions were 30 μm wide × 130 μm wide. An acetone vapor bath was used to smooth acrylonitrile–butadiene–styrene (ABS) surfaces and facilitate bonding of fully enclosed channels. The surfaces of the 3D-printed features were profile...
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3D Printed Micro Free-Flow Electrophoresis Device
2016Co-Authors: Sarah K. Anciaux, Matthew Geiger, Michael T. BowserAbstract:The cost, time, and restrictions on creative flexibility associated with current fabrication methods present significant challenges in the development and application of microfluidic devices. Additive manufacturing, also referred to as three-dimensional (3D) printing, provides many advantages over existing methods. With 3D printing, devices can be made in a cost-effective manner with the ability to rapidly prototype new designs. We have fabricated a micro Free-Flow Electrophoresis (μFFE) device using a low-cost, consumer-grade 3D printer. Test prints were performed to determine the minimum feature sizes that could be reproducibly produced using 3D printing fabrication. Microfluidic ridges could be fabricated with dimensions as small as 20 μm high × 640 μm wide. Minimum valley dimensions were 30 μm wide × 130 μm wide. An acetone vapor bath was used to smooth acrylonitrile–butadiene–styrene (ABS) surfaces and facilitate bonding of fully enclosed channels. The surfaces of the 3D-printed features were profiled and compared to a similar device fabricated in a glass substrate. Stable stream profiles were obtained in a 3D-printed μFFE device. Separations of fluorescent dyes in the 3D-printed device and its glass counterpart were comparable. A μFFE separation of myoglobin and cytochrome c was also demonstrated on a 3D-printed device. Limits of detection for rhodamine 110 were determined to be 2 and 0.3 nM for the 3D-printed and glass devices, respectively
Hainer Wackerbarth - One of the best experts on this subject based on the ideXlab platform.
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Generation of a miniaturized Free-Flow Electrophoresis chip based on a multi-lamination technique—isoelectric focusing of proteins and a single-stranded DNA fragment
Analytical and Bioanalytical Chemistry, 2011Co-Authors: Britta Walowski, Wilhelm Hüttner, Hainer WackerbarthAbstract:Free-Flow Electrophoresis techniques have been applied for separations in various areas of chemistry and biochemistry. Here we focus on the generation of a Free-Flow Electrophoresis chip and direct monitoring of the separation of different molecules in the separation bed of the miniaturized chip. We demonstrate a fast and efficient way to generate a low-cost micro-Free-Flow Electrophoresis (μFFE) chip with a filling capacity of 9.5 μL based on a multi-lamination technique. Separating webs realized by two transfer-adhesive tapes avoid the problem of gas bubbles entering the separation area. The chip is characterized by isoelectric focusing markers (IEF markers). The functionality of the chip is demonstrated by Free-Flow isoelectric focusing (FFIEF) of the proteins BSA (bovine serum albumin) and avidin and a single-stranded DNA (ssDNA) fragment in the pH range 3 to 10. The separation voltage ranges between 167 V cm^−1 and 422 V cm^−1, depending on the application.
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generation of a miniaturized Free Flow Electrophoresis chip based on a multi lamination technique isoelectric focusing of proteins and a single stranded dna fragment
Analytical and Bioanalytical Chemistry, 2011Co-Authors: Britta Walowski, Wilhelm Hüttner, Hainer WackerbarthAbstract:Free-Flow Electrophoresis techniques have been applied for separations in various areas of chemistry and biochemistry. Here we focus on the generation of a Free-Flow Electrophoresis chip and direct monitoring of the separation of different molecules in the separation bed of the miniaturized chip. We demonstrate a fast and efficient way to generate a low-cost micro-Free-Flow Electrophoresis (μFFE) chip with a filling capacity of 9.5 μL based on a multi-lamination technique. Separating webs realized by two transfer-adhesive tapes avoid the problem of gas bubbles enteringthe separation area. The chip is characterized by isoelectric focusing markers (IEF markers). The functionality of the chip is demonstrated by Free-Flow isoelectric focusing (FFIEF) of the proteins BSA (bovine serum albumin) and avidin and a single-stranded DNA (ssDNA) fragment in the pH range 3 to 10. The separation voltage ranges between 167 Vcm −1 and 422 V cm −1 , depending on the application.
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Free-Flow Electrophoresis with electrode-less injection molded chips
Microfluidics BioMEMS and Medical Microsystems IX, 2011Co-Authors: Stefan Köhler, Holger Becker, Volker Beushausen, Erik Beckert, Wilhelm Hüttner, Hainer Wackerbarth, Steffen Howitz, Detlev BelderAbstract:In this work we present the first approach towards low-cost Free-Flow Electrophoresis (FFE) devices utilizing injection molding as a microfabrication process which has the potential to manufacture FFE chips at a cost which make their use commercially viable. This is achieved by realizing a new straightforward micro Free-Flow Electrophoresis (μFFE) design ensuring both, bubble Free electrophoretic separation and effective electrical connection by implementing miniaturized partitioning bars. This creates a defined open gap of 20 μm in height and 500 μm in width between separation zone and electrode channels. The thermoplastic μFFE chips are ready to use, there is no need for a subsequent labor-intensive implementation of membranes or salt bridges to separate the electrode channels from the separation zone.
Christoph Eckerskorn - One of the best experts on this subject based on the ideXlab platform.
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Free Flow Electrophoresis in the proteomic era a technique in flux
Electrophoresis, 2010Co-Authors: Markus Islinger, Christoph Eckerskorn, Alfred VölklAbstract:Since its introduction five decades ago, Free-Flow Electrophoresis (FFE) has been mainly employed for the isolation and fractionation of cells, cell organelles and protein mixtures. In the meantime, the growing interest in the proteome of these bio-particles and biopolymers has shed light on two further facets in the potential of FFE, namely its applicability as an analytical tool and sensor. This review is intended to outline recent innovations, FFE has gained in the proteomic era, and to point out the valuable contributions it has made to the analysis of the proteome of cells, sub-cellular organelles and functional protein networks.
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Free‐Flow Electrophoresis in the proteomic era: A technique in flux
Electrophoresis, 2010Co-Authors: Markus Islinger, Christoph Eckerskorn, Alfred VölklAbstract:Since its introduction five decades ago, Free-Flow Electrophoresis (FFE) has been mainly employed for the isolation and fractionation of cells, cell organelles and protein mixtures. In the meantime, the growing interest in the proteome of these bio-particles and biopolymers has shed light on two further facets in the potential of FFE, namely its applicability as an analytical tool and sensor. This review is intended to outline recent innovations, FFE has gained in the proteomic era, and to point out the valuable contributions it has made to the analysis of the proteome of cells, sub-cellular organelles and functional protein networks.
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Enrichment of Phosphopeptides by Free Flow Electrophoresis
Journal of Proteomics & Bioinformatics, 2008Co-Authors: D. Craft, Gerhard Weber, Christoph Eckerskorn, S. Kronbauer, S. Dower, Craig A. Gelfand, Mikkel NissumAbstract:Protein phosphorylation plays a central role in regulating cellular processes. However, the low level of phosphoproteins in the presence of overwhelming amounts of non-phosphorylated proteins makes their detection and identification challenging. Thus, following tryptic digestion of the proteins, separation of phosphopeptides from non-phosphopeptides is imperative prior to identification. Currently, immobilized metal affinity chromatography (IMAC) and Titanium dioxide supports have been used for phosphorylated peptide enrichment. Gygi et al has also demonstrated the power of using SCX at pH 1.9 to separate phosphorylated peptides from non-phosphorylated peptides successfully. Herein, we present a novel protocol utilizing Free Flow Electrophoresis (FFE) for enrichment and separation of phosphopeptides within complex mixtures.
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Free-Flow Electrophoresis
Cell Biology, 2006Co-Authors: Peter Weber, Gerhard E. Weber, Christoph EckerskornAbstract:Publisher Summary Free-Flow isoetectric focusing of protein mixtures is one of the methods that fulfill the prerequisites to meet the prefractionation demands to increase the amount of low abundant proteins and to dramatically reduce the complexity of protein mixtures. The power and resolution of Free-Flow isoelectric focusing are illustrated with the analysis of pig serum using a Pro Team Free-Flow Electrophoresis (FFE) instrument. One needs to reduce the force of all separation chamber clamps, open the clamps pairwise from the outside to the inside, and open the separation chamber by carefully pulling the front part. Place membranes on electrodes starting from the top to the bottom. The smooth side of the membrane should face toward the electrode seal. The membrane must not protrude over the electrode seal. Subsequently, place the paper strips congruently on the membranes in the same fashion. Place all the media and counterblow tubes in a bottle with distilled water. Place the sample tube in an empty 2-ml reaction tube without tightening the screw.
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Isolation of organelles and prefractionation of protein extracts using Free-Flow Electrophoresis.
Current protocols in protein science, 2004Co-Authors: Peter J A Weber, Gerhard Weber, Christoph EckerskornAbstract:One of the major obstacles in the analysis of proteomes is the extreme complexity of any particular cell or biological fluid. Free-Flow Electrophoresis (FFE) is a powerful tool for reduction of this complexity, which is a prerequisite for systematic and comprehensive protein analyses. Protocols are provided in this unit for sample fractionation at two different stages: on the protein level by isoelectric focusing FFE fractionation of crude protein mixtures such as whole cell lysates, and on a subcellular level by zone-electrophoretic FFE purification of organelles.
