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  • 1.
    Andersson, Olof
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Larsson (Kaiser), Andréas
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Ekblad, Tobias
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Gradient Hydrogel Matrix for Microarray and Biosensor Applications: An Imaging SPR Study2009In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 10, no 1, p. 142-148Article in journal (Refereed)
    Abstract [en]

    A biosensor matrix based on UV-initiated graft copolymerized poly(ethylene glycol) methacrylate and 2-hydroxyethyl methacrylate has been studied using imaging surface plasmon resonance (iSPR). By using a photo mask and a programmable shutter to vary the exposure time laterally, a gradient of matrix spots with physical thicknesses ranging from a few to tens of nanometers was generated. To maximize the dynamic range, imaging SPR was employed in wavelength interrogation mode. By finding the minimum in the reflectance spectra from each pixel of an image, SPR wavelength maps were constructed. The shift in SPR wavelength upon biospecific interaction was then measured both as a function of matrix thickness and composition. The performance of the matrix was evaluated in terms of immobilization of human serum albumin, biomolecular interaction with its antibody, and nonspecific binding of human fibrinogen. In addition, a low molecular weight interaction pair based on a synthetic polypeptide and calmodulin was also studied to explore the size selectivity of the hydrogel matrix. Our results show that the gradient matrix exhibits excellent properties for quick evaluation and screening of optimal hydrogel performance. The mixed hydrogel matrices display very low levels of nonspecific binding. It is also evident that the low molecular weight calmodulin is capable of freely diffusing and interacting throughout the entire hydrogel matrix, whereas the much larger albumin and its corresponding antibody, in particular, are partly/completely hindered from penetrating the interior of the matrix. This size-selectivity is attributed to a significant UV-initiated cross-linking or branching of the matrix during fabrication and/or protein mediated multipoint attachment during immobilization.

  • 2.
    Andersson, Olof
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Larsson (Kaiser), Andréas
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Ekblad, Tobias
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Liedberg, Bo
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Imaging surface plasmon resonance studies of hydrogel and gradient surfaces for biosensor and array applications2008In: Europtrode IX,2008, 2008Conference paper (Other academic)
  • 3.
    Ekblad, Tobias
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Faxälv, Lars
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Chemistry. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Chemistry.
    Andersson, Olof
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Wallmark, Nanny
    Linköping University, Department of Physics, Chemistry and Biology, Molecular Physics. Linköping University, Faculty of Science & Engineering.
    Larsson (Kaiser), Andréas
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Lindahl, Tomas L.
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Chemistry. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Chemistry.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Patterned Hydrogels for Controlled Platelet Adhesion from Whole Blood and Plasma2010In: Advanced Functional Materials, ISSN 1616-301X, E-ISSN 1616-3028, Vol. 20, no 15, p. 2396-2403Article in journal (Refereed)
    Abstract [en]

    This work describes the preparation and properties of hydrogel surface chemistries enabling controlled and well-defined cell adhesion. The hydrogels may be prepared directly on plastic substrates, such as polystyrene slides or dishes, using a quick and experimentally simple photopolymerization process, compatible with photolithographic and microfluidic patterning methods. The intended application for these materials is as substrates for diagnostic cell adhesion assays, particularly for the analysis of human platelet function. The adsorption of fibrinogen and other platelet promoting molecules is shown to be completely inhibited by the hydrogel, provided that the film thickness is sufficient (>5 nm). This allows the hydrogel to be used as a matrix for presenting selected bioactive ligands without risking interference from nonspecifically adsorbed platelet adhesion factors, even in undiluted whole blood and blood plasma. This concept is demonstrated by preparing patterns of proteins on hydrogel surfaces, resulting in highly controlled platelet adhesion. Further insights into the protein immobilization and platelet adhesion processes are provided by studies using imaging surface plasmon resonance. The hydrogel surfaces used in this work appear to provide an ideal platform for cell adhesion studies of platelets, and potentially also for other cell types.

  • 4.
    Larsson (Kaiser), Andréas
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Biochip design based on tailored ethylene glycols2007Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Studies of biomolecular interactions are of interest for several reasons. Beside basic research, the knowledge gained from such studies is also very valuable in for example drug target identification. Medical care is another area where biomolecules may be used as biomarkers to aid physicians in making correct diagnosis. In addition, the highly specific interactions between antibodies and almost any substance opens up the possibilities to design systems for detection of trace amounts of both biological and non-biological substances within environmental restoration, law enforcement, correctional care, customs service and national security. A biochip, which contains a biologically active material, offers a means of monitoring the molecular interactions in the above applications in a sensitive and specific manner. The biochip is a key component of a biosensor, which also includes components for transforming the interaction events into a human-readable signal.

    This thesis describes the use of poly(ethylene glycol) (PEG) in biochip design. Two different approaches are presented, the first based on ethylene glycol (EG)-containing alkyl thiol self-assembled monolayers (SAMs) on flat gold and the second on photo-induced graft copolymerisation of PEG-containing methacrylate monomers onto various substrates. The former is a two dimensional system where EG-terminated thiols are mixed with similar thiols presenting tail groups that mimic the explosive substance 2,4,6-trinitrotoluene (TNT). In an immunoassay, the detection limit for TNT was determined to fall in the range 1-10 µg/L. In the second approach, a branched three dimensional biosensor matrix (hydrogel) is proposed. The carboxymethylated (CM) dextran matrix, which is commonly used within the biosensing community, is not always ideal for studies of biointeractions, due to the non-specific binding frequently encountered in work with complex biological solutions and various proteins. To employ PEG, which displays a low non-specific binding of such species, is therefore an interesting option worth investigating. The use of a branched graft polymerised PEG matrix in biosensor applications is novel as compared to previous reports which have focused on linear PEG chains. The latter approach provides, at maximum, one functional group, per surface anchoring point, for immobilisation of sensor elements. Thus, it has the inherited disadvantage that it limits the number of available immobilisation sites. The present PEG matrix contains a large number of functional groups, for immobilisation of sensor elements, per grafting site and offers the potential of improved response upon binding to the analyte as demonstrated in a series of successful sensor experiments.

    Furthermore, the nature of the process enables easy preparation of matrix patterns and gradients. In a PEG matrix gradient, protein permeability is studied and the capabilities of immobilising proteins are demonstrated. By combining the patterning technique with different monomers in a two-step process, an inert platform, lacking chemical attachment sites, is provided with arrays of spots (with immobilisation capabilities), which are conveniently addressed via microdispensing and used for biosensor purposes. The EG-terminated thiols present another means of generating such inert platforms, a route which is also investigated. To further explore the sensor quality of these spots, the concepts of patterning and gradient formation are combined and studied.

    List of papers
    1. A novel biochip technology for detection of explosives - TNT: Synthesis, characterisation and application
    Open this publication in new window or tab >>A novel biochip technology for detection of explosives - TNT: Synthesis, characterisation and application
    Show others...
    2006 (English)In: Sensors and Actuators B: Chemical, ISSN 0925-4005, Vol. 113, no 2, p. 730-748Article in journal (Refereed) Published
    Abstract [en]

    This contribution describes the synthesis, characterisation and evaluation of a novel biochip technology for the detection of the explosive substance 2,4,6-trinitrotoluene (TNT). Two types of thiols are self-assembled to produce the biochip on gold, namely oligo(ethylene glycol) (OEG)-alkyl thiols terminated with a hydroxyl group and a TNT-analogue (2,4-dinitrobenzene), respectively. Three different TNT-analogues are mixed in various proportions with hydroxyl-terminated OEG-thiols to obtain highly selective and sensitive biochips with a low non-specific binding. The produced self-assembled monolayers (SAMs) are thoroughly characterised with null ellipsometry, contact angle goniometry, infrared reflection absorption spectroscopy (IRAS) and X-ray photoelectron spectroscopy (XPS) and they all meet high standards in terms of molecular conformation, packing and orientation. The biochip is designed to function as a platform for a competitive label-free immunoassay and two real-time transducers – surface plasmon resonance (SPR) and quartz crystal microbalance (QCM) – are used to monitor the dissociation of on-line immobilised monoclonal antibodies produced against TNT. The three TNT-analogues are all potential candidates for the development of a functional biochip, though one of them displayed superior properties in terms of shorter recovery/stabilisation time after antibody immobilisation and a better response/loading capacity ratio. This is particularly evident when using low antigen (TNT-analogue) content in the mixed SAM.

    Keywords
    Explosives; Competitive immunoassay; Self-assembled monolayers; Quartz crystal microbalance; Surface plasmon resonance
    National Category
    Engineering and Technology
    Identifiers
    urn:nbn:se:liu:diva-14605 (URN)10.1016/j.snb.2005.07.025 (DOI)
    Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2015-10-13
    2. Photografted poly(ethylene glycol) matrix for affinity interaction studies
    Open this publication in new window or tab >>Photografted poly(ethylene glycol) matrix for affinity interaction studies
    2007 (English)In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 8, no 1, p. 287-295Article in journal (Refereed) Published
    Abstract [en]

    A poly(ethylene glycol) (PEG)-based matrix for studies of affinity interactions is developed and demonstrated. The PEG matrix, less than 0.1 μm thick, is graft copolymerized onto a cycloolefin polymer from a mixture of PEG methacrylates using a free radical reaction initiated by UV light at 254 nm. The grafting process is monitored in real time, and characteristics such as thickness, homogeneity, relative composition, photostability, and performance in terms of protein resistance in complex biofluids and sensor qualities are investigated with null ellipsometry, infrared spectroscopy, and surface plasmon resonance. The matrix is subsequently modified to contain carboxyl groups, thereby making it possible to immobilize ligands in a controlled and functional manner. Human serum albumin and fibrinogen are immobilized and successfully detected by antibody recognition using surface plasmon resonance. The results are encouraging and suggest that the PEG matrix is suitable for biochip and biosensor applications in demanding biofluids.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-14606 (URN)10.1021/bm060685g (DOI)
    Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2017-12-13
    3. Poly(ethylene glycol) gradient for biochip development
    Open this publication in new window or tab >>Poly(ethylene glycol) gradient for biochip development
    2007 (English)In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 23, no 22, p. 11319-11325Article in journal (Refereed) Published
    Abstract [en]

    A novel method of producing a poly(ethylene glycol) (PEG)-based gradient matrix that varies gradually in thickness from 0 to 500 Å over a distance of 5−20 mm is presented. The gradient matrix is graft copolymerized from a mixture of PEG methacrylates onto organic thin films providing free radical polymerization sites initiated by UV irradiation at 254 nm. The films used as grafting platforms consist of either a spin-coated cycloolefin polymer or a self-assembled monolayer on planar gold. The thickness/irradiation gradient is realized by means of a moving shutter that slowly uncovers the modified gold substrate. The structural and functional characteristics of the gradient matrix are investigated with respect to thickness profile, degree of carboxylation, and subsequent immobilization of two model proteins of different sizes and shapes. These characteristics are studied with ellipsometry and infrared reflection−absorption microscopy using a grazing angle objective. It is revealed that the relatively small carboxylation agent used offers homogeneous activation throughout the gradient, even in the thick areas, whereas the diffusion/interpenetration and subsequent immobilization of large proteins is partially hindered. This is crucial information in biosensor design that can be easily obtained from a gradient experiment on a single sample. Moreover, the partially hindered protein interpenetration, the marginal swelling upon hydration, and the unspecific nature of the graft polymerization suggest a matrix growth mechanism that favors the formation of a bushlike polymer structure with a certain degree of cross linking.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-14607 (URN)10.1021/la700729q (DOI)
    Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2017-12-13
    4. UV-patterned poly(ethylene glycol) matrix for microarray applications
    Open this publication in new window or tab >>UV-patterned poly(ethylene glycol) matrix for microarray applications
    2007 (English)In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 8, no 11, p. 3511-3518Article in journal (Refereed) Published
    Abstract [en]

    A versatile method to fabricate polymeric matrixes for microarray applications is demonstrated. Several different design strategies are presented where a variety of organic films, such as plastic polymers and self-assembled monolayers (SAMs) on planar silica and gold substrates, act as supports for the graft polymerization procedure. An ensemble of poly(ethylene glycol) methacrylate monomers are combined to obtain a matrix with desired properties:  low nonspecific binding and easily accessible groups for postimmobilization of ligands. The free radical graft polymerization process occurs under irradiation with UV light in the 254−266 nm range, which offers the possibility to introduce patterns by means of a photomask. The arrays are created on inert and homogeneous coatings prepared either by graft polymerization of a methoxy-terminated PEG−methacrylate or self-assembly of a methoxy-terminated oligo(ethylene glycol) thiol. Carboxylic acid groups, introduced in the array spots either during graft polymerization or upon wet chemical conversion of hydroxyls, grant the capability to immobilize proteins and other molecules via free amine groups. Immobilization of fluorescent species as well as biotin followed by exposure to a fluorescently labeled antibody directed toward biotin display both excellent integrity of the spots and low nonspecific binding to the surrounding framework. Beside patterns of uniform height and size, an array of spots with varying thickness (a sort of gradient) is demonstrated. Such gradient samples enable us to address critical issues regarding the mechanism(s) behind spatially resolved free radical polymerization of methacrylates. It also offers a convenient route to optimize the matrix properties with respect to thickness, loading capacity, protein diffusion/penetration, and nonspecific binding.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:liu:diva-14608 (URN)10.1021/bm700707s (DOI)
    Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2017-12-13
    5. A hydrogel chip for biosensing studied by imaging surface plasmon resonance
    Open this publication in new window or tab >>A hydrogel chip for biosensing studied by imaging surface plasmon resonance
    Manuscript (Other academic)
    Identifiers
    urn:nbn:se:liu:diva-14609 (URN)
    Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2010-01-13
  • 5.
    Larsson (Kaiser), Andréas
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Angbrant, Johan
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Ekeroth, Johan
    Linköping University, Department of Physics, Chemistry and Biology. Linköping University, The Institute of Technology.
    Månsson, Per
    Biosensor Applications Sweden AB, Sundbyberg, Sweden.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    A novel biochip technology for detection of explosives - TNT: Synthesis, characterisation and application2006In: Sensors and Actuators B: Chemical, ISSN 0925-4005, Vol. 113, no 2, p. 730-748Article in journal (Refereed)
    Abstract [en]

    This contribution describes the synthesis, characterisation and evaluation of a novel biochip technology for the detection of the explosive substance 2,4,6-trinitrotoluene (TNT). Two types of thiols are self-assembled to produce the biochip on gold, namely oligo(ethylene glycol) (OEG)-alkyl thiols terminated with a hydroxyl group and a TNT-analogue (2,4-dinitrobenzene), respectively. Three different TNT-analogues are mixed in various proportions with hydroxyl-terminated OEG-thiols to obtain highly selective and sensitive biochips with a low non-specific binding. The produced self-assembled monolayers (SAMs) are thoroughly characterised with null ellipsometry, contact angle goniometry, infrared reflection absorption spectroscopy (IRAS) and X-ray photoelectron spectroscopy (XPS) and they all meet high standards in terms of molecular conformation, packing and orientation. The biochip is designed to function as a platform for a competitive label-free immunoassay and two real-time transducers – surface plasmon resonance (SPR) and quartz crystal microbalance (QCM) – are used to monitor the dissociation of on-line immobilised monoclonal antibodies produced against TNT. The three TNT-analogues are all potential candidates for the development of a functional biochip, though one of them displayed superior properties in terms of shorter recovery/stabilisation time after antibody immobilisation and a better response/loading capacity ratio. This is particularly evident when using low antigen (TNT-analogue) content in the mixed SAM.

  • 6.
    Larsson (Kaiser), Andréas
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Du, Chun-Xia
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    UV-patterned poly(ethylene glycol) matrix for microarray applications2007In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 8, no 11, p. 3511-3518Article in journal (Refereed)
    Abstract [en]

    A versatile method to fabricate polymeric matrixes for microarray applications is demonstrated. Several different design strategies are presented where a variety of organic films, such as plastic polymers and self-assembled monolayers (SAMs) on planar silica and gold substrates, act as supports for the graft polymerization procedure. An ensemble of poly(ethylene glycol) methacrylate monomers are combined to obtain a matrix with desired properties:  low nonspecific binding and easily accessible groups for postimmobilization of ligands. The free radical graft polymerization process occurs under irradiation with UV light in the 254−266 nm range, which offers the possibility to introduce patterns by means of a photomask. The arrays are created on inert and homogeneous coatings prepared either by graft polymerization of a methoxy-terminated PEG−methacrylate or self-assembly of a methoxy-terminated oligo(ethylene glycol) thiol. Carboxylic acid groups, introduced in the array spots either during graft polymerization or upon wet chemical conversion of hydroxyls, grant the capability to immobilize proteins and other molecules via free amine groups. Immobilization of fluorescent species as well as biotin followed by exposure to a fluorescently labeled antibody directed toward biotin display both excellent integrity of the spots and low nonspecific binding to the surrounding framework. Beside patterns of uniform height and size, an array of spots with varying thickness (a sort of gradient) is demonstrated. Such gradient samples enable us to address critical issues regarding the mechanism(s) behind spatially resolved free radical polymerization of methacrylates. It also offers a convenient route to optimize the matrix properties with respect to thickness, loading capacity, protein diffusion/penetration, and nonspecific binding.

  • 7.
    Larsson (Kaiser), Andréas
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Ekblad, Tobias
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Andersson, Olof
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Photografted poly(ethylene glycol) matrix for affinity interaction studies2007In: Biomacromolecules, ISSN 1525-7797, E-ISSN 1526-4602, Vol. 8, no 1, p. 287-295Article in journal (Refereed)
    Abstract [en]

    A poly(ethylene glycol) (PEG)-based matrix for studies of affinity interactions is developed and demonstrated. The PEG matrix, less than 0.1 μm thick, is graft copolymerized onto a cycloolefin polymer from a mixture of PEG methacrylates using a free radical reaction initiated by UV light at 254 nm. The grafting process is monitored in real time, and characteristics such as thickness, homogeneity, relative composition, photostability, and performance in terms of protein resistance in complex biofluids and sensor qualities are investigated with null ellipsometry, infrared spectroscopy, and surface plasmon resonance. The matrix is subsequently modified to contain carboxyl groups, thereby making it possible to immobilize ligands in a controlled and functional manner. Human serum albumin and fibrinogen are immobilized and successfully detected by antibody recognition using surface plasmon resonance. The results are encouraging and suggest that the PEG matrix is suitable for biochip and biosensor applications in demanding biofluids.

  • 8.
    Larsson (Kaiser), Andréas
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Poly(ethylene glycol) gradient for biochip development2007In: Langmuir, ISSN 0743-7463, E-ISSN 1520-5827, Vol. 23, no 22, p. 11319-11325Article in journal (Refereed)
    Abstract [en]

    A novel method of producing a poly(ethylene glycol) (PEG)-based gradient matrix that varies gradually in thickness from 0 to 500 Å over a distance of 5−20 mm is presented. The gradient matrix is graft copolymerized from a mixture of PEG methacrylates onto organic thin films providing free radical polymerization sites initiated by UV irradiation at 254 nm. The films used as grafting platforms consist of either a spin-coated cycloolefin polymer or a self-assembled monolayer on planar gold. The thickness/irradiation gradient is realized by means of a moving shutter that slowly uncovers the modified gold substrate. The structural and functional characteristics of the gradient matrix are investigated with respect to thickness profile, degree of carboxylation, and subsequent immobilization of two model proteins of different sizes and shapes. These characteristics are studied with ellipsometry and infrared reflection−absorption microscopy using a grazing angle objective. It is revealed that the relatively small carboxylation agent used offers homogeneous activation throughout the gradient, even in the thick areas, whereas the diffusion/interpenetration and subsequent immobilization of large proteins is partially hindered. This is crucial information in biosensor design that can be easily obtained from a gradient experiment on a single sample. Moreover, the partially hindered protein interpenetration, the marginal swelling upon hydration, and the unspecific nature of the graft polymerization suggest a matrix growth mechanism that favors the formation of a bushlike polymer structure with a certain degree of cross linking.

  • 9.
    Liedberg, Bo
    et al.
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Larsson (Kaiser), Andréas
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Ekblad, Tobias
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Andersson, Olof
    Linköping University, The Institute of Technology. Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics.
    Photografted PEG matrix for biosensor applications2006In: Proceedings of the 9th Biosensors Congress,2006, Columbus: American Chemical Society , 2006Conference paper (Refereed)
  • 10.
    Viljanen, Johan
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    Larsson, Jenny
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology. Linköping University, Faculty of Health Sciences.
    Larsson (Kaiser), Andréas
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics. Linköping University, The Institute of Technology.
    Broo, Kerstin S.
    Linköping University, Department of Physics, Chemistry and Biology, Organic Chemistry. Linköping University, The Institute of Technology.
    A Multipurpose Receptor Composed of Promiscuous Proteins. Analyte Detection through Pattern Recognition2007In: Bioconjugate Chemistry, ISSN 1043-1802, Vol. 18, no 6, p. 1935-1945Article in journal (Refereed)
    Abstract [en]

    A multipurpose receptor akin to the “electronic nose” was composed of coumarin-labeled mutants of human glutathione transferase A1. We have previously constructed a kit for site-specific modification of a lysine residue (A216K) using a thiol ester of glutathione (GSC-Coubio) as a modifying reagent. In the present investigation, we scrambled the hydrophobic binding site (H-site) of the protein scaffold through mutations at position M208 via random mutagenesis and isolated a representative library of 11 A216K/M208X mutants. All of the double mutants could be site-specifically labeled to form the K216Cou conjugates. The labeled proteins responded to the addition of different analytes with signature changes in their fluorescence spectra resulting in a matrix of 96 data points per analyte. Ligands as diverse as n-valeric acid, fumaric acid monoethyl ester, lithocholic acid, 1-chloro-2,4-dinitrobenzene (CDNB), glutathione (GSH), S-methyl-GSH, S-hexyl-GSH, and GS-DNB all gave rise to signals that potentially can be interpreted through pattern recognition. The measured Kd values range from low micromolar to low millimolar. The cysteine residue C112 was used to anchor the coumarin-labeled protein to a PEG-based hydrogel chip in order to develop surface-based biosensing systems. We have thus initiated the development of a multipurpose, artificial receptor composed of an array of promiscuous proteins where detection of the analyte occurs through pattern recognition of fluorescence signals. In this system, many relatively poor binders each contribute to detailed readout in a truly egalitarian fashion.

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