Browsing by Subject "Separations"

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    Comprehensive Multidimensional Separations of Biological Samples using Capillary Electrophoresis coupled with Micro Free Flow Electrophoresis
    (2017-12) Johnson, Alexander
    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. The continuous nature of µFFE separations makes it uniquely suitable as the second dimension for multidimensional separations. The focus of this work is the development of coupling capillary electrophoresis (CE) to µFFE as a high speed two-dimensional (2D) separation platform, followed by an investigation of orthogonality of the two techniques, and finally a novel label-free detection method for µFFE separations. A new µFFE device was fabricated and coupled to CE via capillary inserted directly into the µFFE separation channel. High peak capacity separations of trypsin digested BSA and small molecule bioamines demonstrated the power of CE × µFFE. Since both methods rely on electrophoretic mobility to separate, an investigation on the orthogonality of the two techniques was carried out. µFFE can operate in many different separation modes to increase the orthogonality CE × µFFE. Lastly, fluorescent labeling of the analytes can cause the sample to lose its dimensionality affecting 2D separation peak capacity and coverage. A novel absorption detector was studied to demonstrate the first ever label free absorption detection on a µFFE device. A separation was performed on visible dyes and their detection limits quantified.
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    Development, Characterization, and Applications of a 3D Printed micro Free-Flow Electrophoresis Device
    (2017-02) Anciaux, Sarah
    Micro free-flow electrophoresis (μFFE) is a unique separation technique because of its continuous nature. Analytes are pressure driven through a planar separation channel, and an electric field applied laterally to the flow producing a spatial separation. Fabrication methods associated with μFFE devices hinder our ability to address the limitations of μFFE. This work focuses on a novel fabrication method to reduce the overall fabrication cost and time, followed by validating and characterizing the device. A novel μFFE device is fabricated in acrylonitrile butadiene styrene (ABS) by 3D printing two sides of the device and then acetone vapor bonding them while simultaneously inserting electrodes and clarifying the device. Fluorescent dyes are separated, and their limit of detection determined. After validation of the new fabrication method, a new device design is made with the sample inlet modified so that 2D nLC × μFFE separations can be performed. 2D nLC × μFFE separations of fluorescent dyes, proteins, and tryptic BSA digest are demonstrated. These samples allow comparison between the surface properties of glass and 3D printed devices. Peak asymmetries, widths, and the interface were investigated. Minimal surface adsorption is observed for fluorescent dyes, proteins, and peptides, unlike in glass devices. After investigating surface properties, an open edge device for coupling to mass spectrometry is designed and compared to its glass counterpart. A novel ionization method is demonstrated from a hydrophobic membrane and the open edge device is shown to have stable flow.