Robert P. Sebra

University of Colorado at Boulder , Boulder, CO, United States

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Publications (8)27.76 Total impact

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    ABSTRACT: Acrylate monomers were photografted from polymer substrates to create cell responsive chemically and biologically active surfaces that manipulate cell response. Three monomers, polyethylene glycol monoacrylate (MW 375 g/mol) (PEG375A), a monomeric extra-cellular matrix protein, and a cell-cleavable fluorescent monomer, were spatially photopatterned from a base substrate. The base substrate consisted of a dithiocarbamate (DTC) functionalized urethane diacrylate/tri(ethylene glycol)diacrylate copolymer and was shown to non-specifically support NIH 3T3 fibroblast cell adhesion. The DTC-containing polymer was further modified by grafting PEG375A to demonstrate selective blocking of cell-material interactions. Next, acrylated collagen type I was patterned onto polymer substrates to further promote specific cell interactions (i.e. by presenting cell-adhesive moieties). Hydrophilic PEG375A grafted patterns were shown to prevent 3T3 fibroblast adhesion to polymer in spatially grafted regions, while biologically active acrylated collagen type I promoted cell-surface interactions. Collagen type I was grafted at varying densities (0-7.5 pmol/grafted square), and the extent of cell adhesion and spreading were evaluated for each of these graft densities using fluorescence microscopy. Finally, methacrylated carboxyfluorescein diacetate (CFDA) was synthesized and photografted onto a cell-adhesive substrate as a cell sensing mechanism. The acetate groups found in the structure of CFDA cleave in the presence of cells. This cell-responsive substrate results in fluorescence indicative of acetate-group cleavage associated with cell interactions that occurs in patterned regions on polymer surfaces. Collectively, the results herein show the utility and application of a spatially and temporally controlled photografting process for designing cell responsive polymer surfaces.
    Acta Biomaterialia 04/2007; 3(2):151-61. · 5.09 Impact Factor
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    ABSTRACT: Here, we describe the construction of pH sensitive surfaces via the synthesis and controlled photografting of pH sensitive, fluorescent tethers from the surface of a reactive polymeric substrate. The living radical photografting technique presented makes use of dithiocarbamate-functionalized polymer to graft synthetic poly(ethylene glycol) acrylate succinyl fluorescein. Fluorescence intensity of grafted chains is analyzed as a function of photografting reaction time, graft length, buffer solution pH, and cycling sensors from acidic to basic conditions for optical switching. The graft fluorescence response occurs rapidly in a basic environment and grafted functionalities do not cleave or dramatically deplete (up to 72 h later) upon initial exposure to high or low pH buffers. This behavior is a result of the increased stability when introducing the PEG spacer into the structure of the fluorescein. Ultimately, the pH sensitive grafts developed here demonstrate rapid response times, are easy to produce, and are readily integrated onto a fully polymeric microfluidic device using photolithographic techniques and spatially controlled living radical photografting chemistry. Once integrated, sensors such as these could be useful in monitoring pH changes when mixing, reacting, or introducing new chemicals onto a microdevice like the one presented.
    Sensors and Actuators B: Chemical. 11/2006; 119(1):127–134.
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    ABSTRACT: A highly sensitive (pM), efficient (t < 20 min) detection assay was developed by designing surfaces with grafted antibodies. Through this approach, a short half-life antigen, glucagon, was rapidly detected in a biologically complex plasma/blood environment. Tailoring of graft composition eliminates the need for time-consuming blocking steps, significantly reducing antigen-antibody incubation times, while maintaining antibody specificity and activity toward target antigen. Grafted antibodies were bound through solvated, mobile polymer chains, thereby circumventing problems associated with antibody accessibility, analyte diffusion, and steric limitations. The efficiency of this assay is provided through grafting synthesized, acrylated antibodies in the presence of PEG monoacrylate. This procedure eliminates the need for blocking steps, due to a decrease in nonspecific protein interactions. These polymerizable antibodies were tethered with a range of densities while retaining biological activity. Moreover, biological activity of acrylated antibodies was compared to that of unmodified antibodies and remained comparable. The acrylated antibodies were grafted from substrate surfaces using controlled radical photopolymerization, maintaining the advantages of classical antibody immobilization techniques while providing improved detection. Through integrating this antibody conjugation chemistry and immunoassay approach with photolithographic techniques, construction of spatial patterns on a microfluidic device was demonstrated for efficient, parallel screening of multiple antigens.
    Analytical Chemistry 06/2006; 78(9):3144-51. · 5.70 Impact Factor
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    Journal of Polymer Science Part A Polymer Chemistry 01/2006; 44(4):1404 - 1413. · 3.54 Impact Factor
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    ABSTRACT: In this contribution, a new method for the fabrication of complex polymeric microfluidic devices is presented. The technology, contact liquid photolithographic polymerization (CLiPP), overcomes many of the drawbacks associated with other rapid prototyping schemes, such as limited materials choices and time-consuming microassembly protocols. CLiPP shares many traits with other photolithographic methods, but three distinct features: (i) liquid photoresists in contact with the photomask, (ii) readily removed sacrificial materials, and (iii) living radical processes, enable multiple polymeric chemistries and mechanical properties while simultaneously enabling facile fabrication of 3D geometries and surface chemistry control. This contribution details fabrication techniques and methods for the fabrication of high aspect ratio posts covalently bonded to a polymeric substrate, an array of independently stacked bars on top of perpendicular bars, multiple undercut structures fabricated simultaneously, and a complex 3D geometry with intertwined channels.
    Sensors and Actuators B: Chemical. 01/2006;
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    ABSTRACT: The attachment of antibodies to substrate surfaces is useful for achieving specific detection of antigens and toxins associated with clinical and field diagnostics. Here, acrylated whole antibodies were produced through conjugation chemistry, with the goal of covalently photografting these proteins from surfaces in a controlled fashion, to facilitate rapid and sensitive antigenic detection. A living radical photopolymerization chemistry was used to graft the acrylated whole antibodies on polymer surfaces at controlled densities and spatial locations by controlling the exposure time and area, respectively. Copolymer grafts containing these antibodies were synthesized to demonstrate two principles. First, PEG functionalities were introduced to prevent nonspecific protein interactions and improve the reaction kinetics by increasing solvation and mobility of the antibody-containing chains. Both of these properties lead to sensitive (pM) and rapid (<20 min) detection of antigens with this surface modification technique. Second, graft composition was tailored to include multiple antibodies on the same grafted chains, establishing a means for simultaneously detecting multiple antigens on one grafted surface area. Finally, the addition of PEG spacers between the acrylate functionality and the pendant detection antibodies was tuned to enhance the detection of a short-half-life molecule, glucagon, in a complex biological environment, plasma.
    Langmuir 11/2005; 21(24):10907-11. · 4.19 Impact Factor
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    Journal of Polymer Science Part A Polymer Chemistry 04/2005; 43(10):2134 - 2144. · 3.54 Impact Factor
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    ABSTRACT: Microfluidic devices are commonly fabricated in silicon or glass using micromachining technology or elastomers using soft lithography methods; however, invariable bulk material properties, limited surface modification methods and difficulty in fabricating high aspect ratio devices prevent these materials from being utilized in numerous applications and/or lead to high fabrication costs. Contact Liquid Photolithographic Polymerization (CLiPP) was developed as an alternative microfabrication approach that uniquely exploits living radical photopolymerization chemistry to facilitate surface modification of device components, fabrication of high aspect ratio structures from many different materials with numerous covalently-adhered layers and facile construction of three-dimensional devices. This contribution describes CLiPP and demonstrates unique advantages of this new technology for microfabrication of polymeric microdevices. Specifically, the procedure for fabricating devices with CLiPP is presented, the living radical photopolymerization chemistry which enables this technology is described, and examples of devices made using CLiPP are shown.
    Lab on a Chip 01/2005; 4(6):658-62. · 5.70 Impact Factor

Publication Stats

146 Citations
50 Downloads
452 Views
27.76 Total Impact Points

Institutions

  • 2005–2007
    • University of Colorado at Boulder
      • Department of Chemical and Biological Engineering (CHBE)
      Boulder, CO, United States
  • 2006
    • University of Colorado
      Denver, Colorado, United States
  • 2005–2006
    • Howard Hughes Medical Institute
      Maryland, United States