| 1. |
- Trkulja, C. L., et al.
(författare)
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Rational antibody design for undruggable targets using kinetically controlled biomolecular probes
- 2021
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Ingår i: Science Advances. - : American Association for the Advancement of Science (AAAS). - 2375-2548. ; 7:16
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Tidskriftsartikel (refereegranskat)abstract
- Several important drug targets, e.g., ion channels and G protein-coupled receptors, are extremely difficult to approach with current antibody technologies. To address these targets classes, we explored kinetically controlled proteases as structural dynamics-sensitive druggability probes in native-state and disease-relevant proteins. By using low-Reynolds number flows, such that a single or a few protease incisions are made, we could identify antibody binding sites (epitopes) that were translated into short-sequence antigens for antibody production. We obtained molecular-level information of the epitope-paratope region and could produce high-affinity antibodies with programmed pharmacological function against difficult-to-drug targets. We demonstrate the first stimulus-selective monoclonal antibodies targeting the transient receptor potential vanilloid 1 (TRPV1) channel, a clinically validated pain target widely considered undruggable with antibodies, and apoptosis-inducing antibodies selectively mediating cytotoxicity in KRAS-mutated cells. It is our hope that this platform will widen the scope of antibody therapeutics for the benefit of patients.
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| 2. |
- Chiu, D T, et al.
(författare)
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Chemical transformations in individual ultrasmall biomimetic containers
- 1999
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Ingår i: Science. - Stanford Univ, Dept Chem, Stanford, CA 94305 USA. Univ Gothenburg, Dept Chem, S-41296 Gothenburg, Sweden. Pomona Coll, Dept Chem, Claremont, CA 91711 USA. : AMER ASSOC ADVANCEMENT SCIENCE. - 0036-8075 .- 1095-9203. ; 283:5409, s. 1892-1895
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Tidskriftsartikel (refereegranskat)abstract
- Individual phospholipid vesicles, 1 to 5 micrometers in diameter, containing a single reagent or a complete reaction system, were immobilized with an infrared laser optical trap or by adhesion to modified borosilicate glass surfaces. Chemical transformations were initiated either by electroporation or by electrofusion, in each case through application of a short (10-microsecond), intense (20 to 50 kilovolts per centimeter) electric pulse delivered across ultramicroelectrodes. Product formation was monitored by far-field laser fluorescence microscopy. The ultrasmall characteristic of this reaction volume led to rapid diffusional mixing that permits the study of fast chemical kinetics. This technique is also well suited for the study of reaction dynamics of biological molecules within lipid-enclosed nanoenvironments that mimic cell membranes.
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| 3. |
- Chiu, D T, et al.
(författare)
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Electrical and optical methods for the manipulation and analyses of single cells
- 2001
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Ingår i: CLINICAL DIAGNOSTIC SYSTEMS. - Univ Washington, Dept Chem, Seattle, WA 98195 USA. : SPIE-INT SOC OPTICAL ENGINEERING. - 0819439339 ; , s. 1-8
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Konferensbidrag (refereegranskat)abstract
- This paper describes the use of focused electric fields and focused optical fields for the high resolution manipulation of single cells. A focused electric field, obtained with the use of ultramicroelectrodes (tip diameter similar to 5 mum), is used to electroporate and electrofuse individual cells selectively and with high spatial resolution. A focused optical field, in the form of an optical tweezer, is used to isolate single organelles from a cell as well as to position liposomes incorporated with receptors and transporters along the cell for the high-resolution sampling and probing of the cellular microenvironment.
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| 4. |
- Chiu, D T, et al.
(författare)
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Manipulating the biochemical nanoenvironment around single molecules contained within vesicles
- 1999
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Ingår i: Chemical Physics. - Univ Gothenburg, Dept Chem, SE-41296 Gothenburg, Sweden. Stanford Univ, Dept Chem, Stanford, CA 94305 USA. : ELSEVIER. - 0301-0104 .- 1873-4421. ; 247:1, s. 133-139
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Tidskriftsartikel (refereegranskat)abstract
- A method to study single-molecule reactions confined in a biomimetic container is described. The technique combines rapid vesicle preparation, optical trapping and fluorescence confocal microscopy for performing simultaneous single-vesicle trapping and single-molecule detection experiments. The collisional environment between a single enzyme and substrate inside a vesicle is characterized by a Brownian dynamics Monte Carlo simulation. (C) 1999 Elsevier Science B.V. All rights reserved.
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| 5. |
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| 6. |
- Jeffries, GDM, et al.
(författare)
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3D micro-organisation printing of mammalian cells to generate biological tissues
- 2020
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Ingår i: Scientific reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 10:1, s. 19529-
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Tidskriftsartikel (refereegranskat)abstract
- Significant strides have been made in the development of in vitro systems for disease modelling. However, the requirement of microenvironment control has placed limitations on the generation of relevant models. Herein, we present a biological tissue printing approach that employs open-volume microfluidics to position individual cells in complex 2D and 3D patterns, as well as in single cell arrays. The variety of bioprinted cell types employed, including skin epithelial (HaCaT), skin cancer (A431), liver cancer (Hep G2), and fibroblast (3T3-J2) cells, all of which exhibited excellent viability and survivability, allowing printed structures to rapidly develop into confluent tissues. To demonstrate a simple 2D oncology model, A431 and HaCaT cells were printed and grown into tissues. Furthermore, a basic skin model was established to probe drug response. 3D tissue formation was demonstrated by co-printing Hep G2 and 3T3-J2 cells onto an established fibroblast layer, the functionality of which was probed by measuring albumin production, and was found to be higher in comparison to both 2D and monoculture approaches. Bioprinting of primary cells was tested using acutely isolated primary rat dorsal root ganglia neurons, which survived and established processes. The presented technique offers a novel open-volume microfluidics approach to bioprint cells for the generation of biological tissues.
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| 7. |
- Karlsson, M, et al.
(författare)
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Electroinjection of colloid particles and biopolymers into single unilamellar liposomes and cells for bioanalytical applications
- 2000
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Ingår i: Analytical Chemistry. - Univ Gothenburg, Dept Chem, SE-41296 Gothenburg, Sweden. Univ Gothenburg, Dept Chem Phys, SE-41345 Gothenburg, Sweden. : AMER CHEMICAL SOC. - 0003-2700 .- 1520-6882. ; 72:23, s. 5857-5862
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Tidskriftsartikel (refereegranskat)abstract
- A combined electroporation and pressure-driven microinjection method for efficient loading of biopolymers and colloidal particles into single-cell-sized unilamellar liposomes was developed. Single liposomes were positioned between a similar to2-mum tip diameter solute-filled glass micropipet, equipped with a Pt electrode, and a 5-mum-diameter carbon fiber electrode. A transient, 1-10 ms, rectangular waveform de voltage pulse (10-40 V/cm) was applied between the electrodes, thus focusing the electric field over the liposome. Dielectric membrane breakdown induced by the applied voltage pulse caused the micropipet tip to enter the liposome and a small volume (typically 50-500 x 10(-15) L) of fluorescein, YOYO-intercalated T7-phage DNA, 100-nm-diameter unilamellar liposomes, or fluorescent latex spheres could be injected into the intraliposomal compartment. We also demonstrate initiation of a chemical intercalation reaction between T2-phage DNA and YOYO-1 by dual injection into a single giant unilamellar liposome. The method was also successfully applied for loading of single cultured cells.
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| 8. |
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| 9. |
- Strömberg, Anette, et al.
(författare)
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Manipulating the genetic identity and biochemical surface properties of individual cells with electric-field-induced fusion
- 2000
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - Stanford Univ, Dept Chem, Stanford, CA 94305 USA. Gothenburg Univ, Dept Chem, S-41296 Gothenburg, Sweden. Sahlgrens Univ Hosp, Inst Clin Neurosci, Dept Neurol, S-41345 Gothenburg, Sweden. : NATL ACAD SCIENCES. - 0027-8424 .- 1091-6490. ; 97:1, s. 7-11
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Tidskriftsartikel (refereegranskat)abstract
- A method for cell-cell and cell-liposome fusion at the single-cell level is described. Individual cells or liposomes were first selected and manipulated either by optical trapping or by adhesion to a micromanipulator-controlled ultramicroelectrode. Spatially selective fusion of the cell-cell or cell-liposome pair was achieved by the application of a highly focused electric field through a pair of 5-mu m o.d. carbon-fiber ultramicroelectrodes. The ability to fuse together single cells opens new possibilities in the manipulation of the genetic and cellular makeup of individual cells in a controlled manner, In the study of cellular networks, for example, the alteration of the biochemical identity of a selected cell can have a profound effect on the behavior of the entire network. Fusion of a single liposome with a target cell allows the introduction of the liposomal content into the cell interior as well as the addition of lipids and membrane proteins onto the cell surface. This cell-liposome fusion represents an approach to the manipulation of the cytoplasmic contents and surface properties of single cells. As an example, we have introduced a membrane protein (gamma-glutamyltransferase) reconstituted in liposomes into the cell plasma membrane.
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| 10. |
- Strömberg, Anette, et al.
(författare)
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Microfluidic device for combinatorial fusion of liposomes and cells
- 2001
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Ingår i: Analytical Chemistry. - Gothenburg Univ, Dept Chem, S-41296 Gothenburg, Sweden. : AMER CHEMICAL SOC. - 0003-2700 .- 1520-6882. ; 73:1, s. 126-130
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Tidskriftsartikel (refereegranskat)abstract
- We describe an electrofusion-based technique for combinatorial synthesis of individual liposomes. A prototype device with containers for liposomes of different compositions and a fusion container was constructed. The sample containers had fluid contact with the fusion container through microchannels. Optical trapping was used to transport individual liposomes and cells through the microchannels into the fusion container. In the fusion container, selected pairs of liposomes were fused together using microelectrodes. A large number of combinatorially synthesized Liposomes with complex compositions and reaction systems can be obtained from small sets of precursor liposomes. The order of different reaction steps can be specified and defined by the fusion sequence. This device could also facilitate single cell-cell electrofusions (hybridoma production). This is exemplified by fusion of transported red blood cells.
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| 11. |
- Wilson, C F, et al.
(författare)
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Nanoengineered structures for holding and manipulating liposomes and cells
- 2001
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Ingår i: Analytical Chemistry. - Stanford Univ, Dept Chem, Stanford, CA 94305 USA. Univ Gothenburg, Dept Chem, SE-41296 Gothenburg, Sweden. : AMER CHEMICAL SOC. - 0003-2700 .- 1520-6882. ; 73:4, s. 787-791
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Tidskriftsartikel (refereegranskat)abstract
- We describe the fabrication of nanoengineered holding pipets with concave seating surfaces and fine pressure control. These pipets were shown to exhibit exceptional stability in capturing, transporting, and releasing single cells and liposomes 1-12 mum in diameter, which opens previously inaccessible avenues of research. Three specific examples demonstrated the, utility and versatility of this manipulation system. In the first, carboxyrhodaminie was selectively incorporated into individual cells by electroporation, after which nearly all the medium (hundreds of microliters) surrounding the docked and tagged cells was rapidly exchanged (in seconds) and the cells were subsequently probed by laser-induced fluorescence (LIF). In the second study, a single liposome containing carboxyrhodamine was transported to a dye-free solution using a transfer pipet, docked to a holding pipet, and held firmly during physical agitation and interrogation by LIF. In the third study, pairs of liposomes were positioned between two microelectrodes, held in contact, and selectively electrofused and the resulting liposomes undocked intact.
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