| 1. |
- Cans, Ann-Sofie, 1971, et al.
(författare)
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A Secretory Artificial Cell for Exocytosis
- 2013
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Ingår i: Biophysical Journal. - : Elsevier BV. - 0006-3495 .- 1542-0086. ; 104:2 Suppl 1
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- The complexity of exocytosis has left the molecular details of the process unclear. We present a minimal, artificial secretory cell designed for amperometric studies of release of signalling molecules through the fusion pore of single vesicles. In replacement of SNARE-proteins, the cell model has been equipped with an analog composed of complimentary DNA constructs, one on the vesicle and one on the target membrane. The DNA constructs hybridize in a zipper-like fashion bringing about docking of the vesicles and following the addition of Ca2+, fusion of the vesicles is completed. Exocytotic events recorded from the artificial cell closely approximate exocytosis in live cells. The results together with simulations of vesicular release demonstrate that the lipid-based fusion pore initially retains stability and limits diffusion of the secreted molecules.
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| 2. |
- Simonsson, Lisa, 1982, et al.
(författare)
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A functioning artificial secretory cell
- 2012
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322 .- 2045-2322. ; 2, s. no. 824-
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Tidskriftsartikel (refereegranskat)abstract
- We present an amperometric study of content release from individual vesicles in an artificial secretory cell designed with the minimal components required to carry out exocytosis. Here, the membranes of the cell and vesicles are substituted for protein-free giant and large unilamellar vesicles respectively. In replacement of the SNARE-complex, the cell model was equipped with an analog composed of complimentary DNA constructs. The DNA constructs hybridize in a zipper-like fashion to bring about docking of the artificial secretory vesicles and following the addition of Ca2+ artificial exocytosis was completed. Exocytotic events recorded from the artificial cell closely approximate exocytosis in live cells. The results together with simulations of vesicular release demonstrate that the molecular flux in this model is attenuated and we suggest that this is the result of restricted diffusion through a semi-stable fusion pore or a partitioning of the signalling molecule out of the fused vesicle membrane.
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| 3. |
- Harreither, Wolfgang, et al.
(författare)
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Carbon Nanotube Fiber Microelectrodes Show a Higher Resistance to Dopamine Fouling
- 2013
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 85:15, s. 7447-7453
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Tidskriftsartikel (refereegranskat)abstract
- We have compared the properties and resistance to DA fouling of a carbon nanotube fiber (CNTF) microelectrode to a traditional carbon fiber (CF) microelectrode. These two materials show comparable electrochemical activities for outer-sphere and inner-sphere redox reactions. Although the CNTF might have a higher intrinsic RC constant, thus limiting its high-frequency behavior, the CNTF shows a significantly higher durability than the CF in terms of electrode stability. During constant oxidation of 100 mu M DA, the signal measured by the CNTF rnicroelectrode shows a 2-h window over which no decrease in current is observed. Under the same conditions, the current obtained at the CF microelectrode decreases by almost 50%. A model of the fouling process, assuming the formation of growing patches of insulator on the surface, has been compared to the data. This model is found to be in good agreement with our results and indicates a growth rate of the patches in the 0.1-2 nm s(-1) range.
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| 4. |
- Harreither, Wolfgang, et al.
(författare)
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Cysteine residues reduce the severity of dopamine electrochemical fouling
- 2016
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Ingår i: Electrochimica Acta. - : Elsevier BV. - 0013-4686. ; 210, s. 622-629
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Tidskriftsartikel (refereegranskat)abstract
- The fouling of neurotransmitter during electrochemical detection can dramatically jeopardize the viability of the sensor. Several recent developments in electrode fabrication and design have helped mitigating this issue. For instance, carbon nanotube fiber (CNTF) electrodes were found to be more resistant to fouling than the traditional carbon fiber (CF) electrodes. Here, we investigate the fouling resistance of these two materials in the presence of albumin. Interestingly, our data shows that the presence of albumin reduces the impact of dopamine (DA) fouling in a dose-dependent manner. A protective effect from DA fouling was also observed for other thiol containing substances, thus hinting that the sulfur competes with the amine moiety for the nucleophilic binding to the oxidized catechol, a critical step initiating DA fouling, and therefore reduces the rate of DA polymerization on the electrode surface. Overall, this study furthers our understanding of the mechanisms controlling DA detection in situ, and suggests using solutions of albumin as a background for calibrating sensors.
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| 5. |
- Lin, Yuqing, 1980, et al.
(författare)
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Chemical Analysis of Single Cells
- 2011
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 1520-6882 .- 0003-2700. ; 83:12, s. 4369-4392
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Tidskriftsartikel (refereegranskat)
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| 6. |
- Mellander, Lisa J., et al.
(författare)
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Amperometric post spike feet reveal most exocytosis is via extended kiss-and-run fusion
- 2012
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 2:artikel nr 907, s. 1-6
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Tidskriftsartikel (refereegranskat)abstract
- The basis for communication between nerve cells lies in the process of exocytosis, the fusion of neurotransmitter filled vesicles with the cell membrane resulting in release of the signaling molecules. Even though much is known about this process, the extent that the vesicles are emptied upon fusion is a topic that is being debated. We have analyzed amperometric peaks corresponding to release at PC12 cells and find stable plateau currents during the decay of peaks, indicating closing of the vesicle after incomplete release of its content. Using lipid incubations to alter the amount of transmitter released we were able to estimate the initial vesicular content, and from that, the fraction of release. We propose a process for most exocytosis events where the vesicle partially opens to release transmitter and then closes directly again, leaving the possibility for regulation of transmission within events.
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| 7. |
- Passarelli, Melissa, 1983, et al.
(författare)
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Development of an Organic Lateral Resolution Test Device for Imaging Mass Spectrometry
- 2014
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 86:19, s. 9473-9480
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Tidskriftsartikel (refereegranskat)abstract
- An organic lateral resolution test device has been developed to measure the performance of imaging mass spectrometry (IMS) systems. The device contains periodic gratings of polyethylene glycol (PEG) and lipid bars covering a wide range of spatial frequencies. Microfabrication technologies were employed to produce well-defined chemical interfaces, which allow lateral resolution to be assessed using the edge-spread function (ESF). In addition, the design of the device allows for the direct measurement of the modulation transfer function (MTF) to assess image quality. Scanning electron microscopy (SEM) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) were used to characterize the device. TOF-SIMS imaging was used to measure the chemical displacement of biomolecules in matrix-assisted laser desorption/ionization (MALDI) matrix crystals. In a proof-of-concept experiment, the platform was also used to evaluate MALDI matrix application methods, specifically aerosol spray and sublimation methods.
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| 8. |
- Trouillon, Raphaël, 1982, et al.
(författare)
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Actin Controls the Vesicular Fraction of Dopamine Released During Extended Kiss and Run Exocytosis
- 2014
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Ingår i: Acs Chemical Biology. - : American Chemical Society (ACS). - 1554-8929 .- 1554-8937. ; 9:3, s. 812-820
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Tidskriftsartikel (refereegranskat)abstract
- The effect of latrunculin A, an inhibitor of actin cross-linking, on exocytosis in PC12 cells was investigated with single cell amperometry. This analysis strongly suggests that the actin cytoskeleton might be involved in regulating exocytosis, especially by mediating the constriction of the pore. In an extended kiss-and-run release mode, actin could actually control the fraction of neurotransmitters released by the vesicle. This scaffold appears to contribute, with the lipid membrane and the protein machinery, to the closing dynamics of the pore, in competition with other forces mediating the opening of the exocytotic channel.
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| 9. |
- Trouillon, Raphaël, 1982, et al.
(författare)
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Amperometric Measurements at Cells Support a Role for Dynamin in the Dilation of the Fusion Pore during Exocytosis
- 2013
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Ingår i: Chemphyschem. - : Wiley. - 1439-4235 .- 1439-7641. ; 14:10, s. 2295-2301
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Tidskriftsartikel (refereegranskat)abstract
- Dynamin is a GTPase mechanochemical enzyme involved in the late steps of endocytosis, where it separates the endocytotic vesicle from the cell membrane. However, recent reports have emphasized its role in exocytosis. In this case, dynamin may contribute to the control of the exocytotic pore, thus suggesting a direct control on the efflux of neurotransmitters. Dynasore, a selective inhibitor of the GTPase activity of dynamin, was used to investigate the role of dynamin in exocytosis. Exocytosis was analyzed by amperometry, thus revealing that dynasore inhibits exocytosis in a dose-dependent manner. Analysis of the exocytotic peaks shows that the inhibition of the GTPase activity of dynamin leads to shorter, smaller events. This observation, together with the rapid effect of dynasore, suggests that the blocking of the GTPase induces the formation of a more narrow and short-lived fusion pore. These results suggest that the GTPase properties of dynamin are involved in the duration and kinetics of exocytotic release. Interestingly, and in strong contrast with its role in endocytosis, the mechanochemical properties of dynamin appear to contribute to the dilation and stability of the pore during exocytosis.
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| 10. |
- Trouillon, Raphaël, 1982, et al.
(författare)
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An electrochemical functional assay for the sensing of nitric oxide release induced by angiogenic factors
- 2011
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Ingår i: BMB REPORTS. - 1976-6696. ; 44:11, s. 699-704
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Forskningsöversikt (refereegranskat)abstract
- Abstract: Nitric oxide (NO) is a critical biological mediator involved in numerous diseases. However, the short lifetime of this molecule in biological conditions can make its study in situ complicated. Here, we review some recent results on the role of NO in angiogenesis, obtained using a biocompatible micro-electrode array. This simple system allowed for the quick and easy quantification of NO released from cells grown directly on the surface of the sensor. We have used this technology to demonstrate that angiogenin induces NO release, and to partially elucidate its intracellular transduction pathway. [BMB reports 2011; 44(11): 699-704]
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