| 1. |
- Klionsky, Daniel J., et al.
(författare)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Forskningsöversikt (refereegranskat)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
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| 2. |
- Aad, G., et al.
(författare)
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- 2012
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swepub:Mat__t (refereegranskat)
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| 3. |
- Aad, G., et al.
(författare)
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- 2011
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swepub:Mat__t (refereegranskat)
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| 4. |
- Hsu, Ya-Ching, et al.
(författare)
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Methadone concentrations in blood, plasma, and oral fluid determined by isotope-dilution gas chromatography-mass spectrometry
- 2013
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Verlag (Germany). - 1618-2642 .- 1618-2650. ; 405:12, s. 3921-3928
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Tidskriftsartikel (refereegranskat)abstract
- Methadone (MTD) is widely used for detoxification of heroin addicts and also in pain management programs. Information about the distribution of methadone between blood, plasma, and alternative specimens, such as oral fluid (OF), is needed in clinical, forensic, and traffic medicine when analytical results are interpreted. We determined MTD and its metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) in blood, plasma, blood cells, and OF by gas chromatography-mass spectrometry (GC-MS) after adding deuterium-labeled internal standards. The analytical limits of quantitation for MTD and EDDP by this method were 20 and 3 ng/mL, respectively. The amounts of MTD and EDDP were higher in plasma (80.4 % and 76.5 %) compared with blood cells (19.6 % and 23.5 %) and we found that repeated washing of blood cells with phosphate-buffered saline increased the amounts in plasma (93.6 % and 88.6 %). Mean plasma/blood concentration ratios of MTD and EDDP in spiked samples (N = 5) were 1.27 and 1.21, respectively. In clinical samples from patients (N = 46), the concentrations of MTD in plasma and whole blood were highly correlated (r = 0.92, p andlt; 0.001) and mean (median) plasma/blood distribution ratios were 1.43 (1.41). The correlations between MTD in OF and plasma (r = 0.46) and OF and blood (r = 0.52) were also statistically significant (p andlt; 0.001) and the mean OF/plasma and OF/blood distribution ratios were 0.55 and 0.77, respectively. The MTD concentration in OF decreased as salivary pH increased (more basic). These results will prove useful in clinical and forensic medicine when MTD concentrations in alternative specimens are compared and contrasted.
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| 6. |
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| 7. |
- Chen, Li-li, et al.
(författare)
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Multilayered polyelectrolyte-coated gold nanorods as multifunctional optical contrast agents for cancer cell imaging
- 2010
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Ingår i: Journal of Zhejiang University: Science B. - 1673-1581. ; 11:6, s. 417-422
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Tidskriftsartikel (refereegranskat)abstract
- We report the application of multilayered polyelectrolyte-coated gold nanorods (GNRs) as multifunctional optical contrast agents for cancer cell imaging. The surface modification of GNRs improves their chemical stability and facilitates them to be taken up by cancer cells through electrostatic interaction. The unique longitudinal surface plasmon resonance property of GNRs makes them suitable as both "scattering contrast agents" and "Raman contrast agents". In our experiments, the staining of GNRs in cells was further confirmed by dark field microscopy and Raman microscopy. Our experiment results indicated that GNRs have great potential as multifunctional "optical contrast agents" for future in vivo animal imaging.
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| 8. |
- Chen, Shu-Feng, et al.
(författare)
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Chemiluminescence of Coelenterazine and Fluorescence of Coelenteramide : A Systematic Theoretical Study
- 2012
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Ingår i: Journal of Chemical Theory and Computation. - : American Chemical Society (ACS). - 1549-9618 .- 1549-9626. ; 8:8, s. 2796-2807
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Tidskriftsartikel (refereegranskat)abstract
- A systematic investigation of the structural and spectroscopic properties of coelenteramide has been performed at the TD-CAM-B3LYP/6-31+G(d,p) level of theory, including various fluorescence and chemiluminescence states. The influence of geometric conformations, solvent polarity, protonation state, and the covalent character of the O-H bond of the hydroxyphenyl moiety were carefully studied. Striking differences in geometries and electronic structures among the states responsible for light emission were characterized. All fluorescence states can be described as a limited charge transfer process for a planar amide moiety. However, the chemiluminescence state is characterized by a much larger charge transfer that takes place over a longer distance. Moreover, the chemiluminescent coelenteramide structure exhibits an amide moiety that is no longer planar, in agreement with recent, more accurate ab initio results [Roca-Sanjuan et al J. Chem. Theory Comput. 2011, 7, 4060] Because the chemiluminescence state appears to be completely dark, a new mechanism is tentatively introduced for this process.
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| 9. |
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| 10. |
- Chen, Shu-Feng, et al.
(författare)
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Multireference theoretical studies on the solvent effect of firefly multicolor bioluminescence
- 2011
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Ingår i: International Journal of Quantum Chemistry. - : Wiley. - 0020-7608 .- 1097-461X. ; 111:13, s. 3371-3377
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Tidskriftsartikel (refereegranskat)abstract
- In concert with the recent spectroscopic studies of the light-color modulation mechanism of firefly (Hirano et al., J Am Chem Soc 2009, 131, 2385), quantum chemical calculations using complete active space SCF (CASSCF), multistate complete active space second order perturbation (MS-CASPT2) theory as well as a time-dependent density functional theory (TD-DFT) approach with the Coulomb attenuated hybrid exchange-correlation functional (CAM-B3LYP) were performed on the excited state (S1) of the keto-form oxyluciferin (keto-OxyLH2). Benzene, DMSO, CH3CN, and H2O were chosen as polar solvents. The polarization effect of less polar solvent was considered by a simple model, complex of keto-OxyLH2, and NH3 with different covalent character. The calculated results supported the experimental conclusion: (1) the light emitter of bioluminescence is the S1 state of keto-OxyLH2 anion [(keto-1)*], and (2) light emission from (keto-1)* is modulated by the polarity of surrounding environment and the degree of covalent character of hydrogen bond between (keto-1)* and a protonated basic moiety. The mechanism of the multicolor bioluminescence was discussed from the theoretical viewpoint.
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