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- 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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- 2019
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Tidskriftsartikel (refereegranskat)
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- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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- Kanoni, Stavroula, et al.
(författare)
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Implicating genes, pleiotropy, and sexual dimorphism at blood lipid loci through multi-ancestry meta-analysis.
- 2022
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Ingår i: Genome biology. - : Springer Science and Business Media LLC. - 1474-760X .- 1465-6906 .- 1474-7596. ; 23:1
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Tidskriftsartikel (refereegranskat)abstract
- Genetic variants within nearly 1000 loci are known to contribute to modulation of blood lipid levels. However, the biological pathways underlying these associations are frequently unknown, limiting understanding of these findings and hindering downstream translational efforts such as drug target discovery.To expand our understanding of the underlying biological pathways and mechanisms controlling blood lipid levels, we leverage a large multi-ancestry meta-analysis (N=1,654,960) of blood lipids to prioritize putative causal genes for 2286 lipid associations using six gene prediction approaches. Using phenome-wide association (PheWAS) scans, we identify relationships of genetically predicted lipid levels to other diseases and conditions. We confirm known pleiotropic associations with cardiovascular phenotypes and determine novel associations, notably with cholelithiasis risk. We perform sex-stratified GWAS meta-analysis of lipid levels and show that 3-5% of autosomal lipid-associated loci demonstrate sex-biased effects. Finally, we report 21 novel lipid loci identified on the X chromosome. Many of the sex-biased autosomal and X chromosome lipid loci show pleiotropic associations with sex hormones, emphasizing the role of hormone regulation in lipid metabolism.Taken together, our findings provide insights into the biological mechanisms through which associated variants lead to altered lipid levels and potentially cardiovascular disease risk.
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- Ablikim, M., et al.
(författare)
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Amplitude analysis of the pi(0)pi(0) system produced in radiative J/psi decays
- 2015
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Ingår i: Physical Review D. - 1550-7998 .- 1550-2368. ; 92:5
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Tidskriftsartikel (refereegranskat)abstract
- An amplitude analysis of the pi(0)pi(0) system produced in radiative J/psi decays is presented. In particular, a piecewise function that describes the dynamics of the pi(0)pi(0) system is determined as a function of M pi(0)pi(0) from an analysis of the (1.311 +/- 0.011) x 10(9) J/psi decays collected by the BESIII detector. The goal of this analysis is to provide a description of the scalar and tensor components of the pi(0)pi(0) system while making minimal assumptions about the properties or number of poles in the amplitude. Such a model-independent description allows one to integrate these results with other related results from complementary reactions in the development of phenomenological models, which can then be used to directly fit experimental data to obtain parameters of interest. The branching fraction of J/psi -> pi(0)pi(0) is determined to be (1.15 +/- 0.05) x 10(-3), where the uncertainty is systematic only and the statistical uncertainty is negligible.
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- Ablikim, M., et al.
(författare)
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An improved limit for Gamma(ee) of X(3872) and Gamma(ee) measurement of psi(3686)
- 2015
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Ingår i: Physics Letters B. - : Elsevier BV. - 0370-2693 .- 1873-2445. ; 749, s. 414-420
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Tidskriftsartikel (refereegranskat)abstract
- Using the data sets taken at center-of-mass energies above 4 GeV by the BESIII detector at the BEPCII storage ring, we search for the reaction e(+)e(-) -> gamma(ISR) X(3872) -> gamma(ISR)pi(+)pi(-) J/psi via the Initial State Radiation technique. The production of a resonance with quantum numbers J(PC) = 1(++) such as the X(3872) via single photon e(+)e(-) annihilation is forbidden, but is allowed by a next-to-leading order box diagram. We do not observe a significant signal of X(3872), and therefore give an upper limit for the electronic width times the branching fraction Gamma B-X(3872)(ee)(X(3872) -> pi(+)pi(-) J/psi) < 0.13 eVat the 90% confidence level. This measurement improves upon existing limits by a factor of 46. Using the same final state, we also measure the electronic width of the psi(3686) to be Gamma(psi)(ee)(3686) ee = 2213 +/- 18(stat) +/- 99(sys) eV.
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- Ablikim, M., et al.
(författare)
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Branching fraction measurement of J/ψ→KSKL and search for J/ψ→KSKS
- 2017
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Ingår i: Physical Review D. - 2470-0010 .- 2470-0029. ; 96:11
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Tidskriftsartikel (refereegranskat)abstract
- Using a sample of 1.31 x 10(9) J/Psi events collected with the BESIII detector at the BEPCII collider, we study the decays of J/Psi -> KSKL and KSKS. The branching fraction of J/Psi -> KSKL is determined to be B(J/Psi -> KSKL) = (1.93 +/- 0.01 (stat) +/- 0.05 (syst)) x 10(-4), which significantly improves on previous measurements. No clear signal is observed for the J/Psi -> KSKS process, and the upper limit at the 95% confidence level for its branching fraction is determined to be B(J/Psi -> KSKS) < 1.4 x 10(-8), which improves on the previous searches by 2 orders in magnitude and reaches the order of the Einstein-Podolsky-Rosen expectation.
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