| 1. |
- Beal, Jacob, et al.
(författare)
-
Robust estimation of bacterial cell count from optical density
- 2020
-
Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
-
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.
|
|
| 2. |
- Klionsky, Daniel J., et al.
(författare)
-
Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
-
Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
-
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.
|
|
| 3. |
- Liu, Chang, et al.
(författare)
-
Oligomer Dynamics of LL-37 Truncated Fragments Probed by α-Hemolysin Pore and Molecular Simulations
- 2023
-
Ingår i: Small. - 1613-6810 .- 1613-6829. ; 19:37
-
Tidskriftsartikel (refereegranskat)abstract
- Oligomerization of antimicrobial peptides (AMPs) is critical in their effects on pathogens. LL-37 and its truncated fragments are widely investigated regarding their structures, antimicrobial activities, and application, such as developing new antibiotics. Due to the small size and weak intermolecular interactions of LL-37 fragments, it is still elusive to establish the relationship between oligomeric states and antimicrobial activities. Here, an α-hemolysin nanopore, mass spectrometry (MS), and molecular dynamic (MD) simulations are used to characterize the oligomeric states of two LL-37 fragments. Nanopore studies provide evidence of trapping events related to the oligomer formation and provide further details on their stabilities, which are confirmed by MS and MD simulations. Furthermore, simulation results reveal the molecular basis of oligomer dynamics and states of LL-37 fragments. This work provides unique insights into the relationship between the oligomer dynamics of AMPs and their antimicrobial activities at the single-molecule level. The study demonstrates how integrating methods allows deciphering single molecule level understanding from nanopore sensing approaches.
|
|
| 4. |
- Liu, Yang, et al.
(författare)
-
Evolution of residual stress when turning a fillet radius in stainless steel
- 2023
-
Ingår i: Journal of Manufacturing Processes. - : Elsevier BV. - 1526-6125. ; 85, s. 216-226
-
Tidskriftsartikel (refereegranskat)abstract
- Most studies have been carried out to investigate the surface integrity induced by metal cutting process. However, the previous studies are limited to a longitudinal turning or orthogonal cutting operations and the residual stresses generated in a fillet radius have been ignored. This study uses a combination of experiments and numerical simulations to study the evolution of cutting forces, temperature, chip morphology, and residual stress distributions while turning a fillet radius in AISI 304. Finite Element (FE) models were developed with a Coupled Eulerian and Lagrangian (CEL) method, where the geometric model of the workpiece was established taking into account the previous machined surface profile at the four specific cutting faces. The model was validated by experimental cutting forces, chip morphology, and residual stress profiles. The changing trend of shape and area of uncut chip cross-section during fillet turning were analyzed to explain the evolution of cutting forces and temperatures. The results show that the cutting force components in cutting speed and tangential directions increase during the early stage of the fillet turning process and decrease after that, while the force in the radial direction shows an increasing trend during this process. The maximum temperature at the machined surface is increased along the tool path. In addition, magnitude and depth of residual stress are slightly changed during the fillet radius turning process, but a reduction of the residual stress profile can still be noticed.
|
|
| 5. |
- Liu, Yang, et al.
(författare)
-
FE analysis on the association between tool edge radius and thermal-mechanical load in machining Inconel 718
- 2021
-
Ingår i: 18th CIRP Conference on Modeling of Machining Operations (CMMO), Ljubljana, Slovenia, June 15-17, 2021. - : Elsevier BV. - 2212-8271. ; 102, s. 91-96
-
Konferensbidrag (refereegranskat)abstract
- The cutting edge geometry has a significant influence on thermal-mechanical loads and the quality of machined surface in machining operations. In this study, a joint numerical and experimental investigation was conducted to study the effects of tool edge radius on process variables (tool temperatures, force components, stresses) and process dynamics in orthogonal machining Inconel 718. The numerical model was implemented based on Coupled Eulerian-Lagrangian (CEL) formulation, in which the tool is regarded as elastic, making it possible to realise the fluctuation of the tool tip. The results show that the temperature at the tool tip is more sensitive to the change of edge radius than the maximum temperature at the tool-chip interface. The force components acting on rake face and flank face increase with increasing edge radius due to more negative effective rake angle and increasing ploughing depth respectively. Besides, the generated chip tends to be more segmented with increase of edge radius, causing severer fluctuation of forces and tool tip displacement. It was also found that a larger edge radius leads to a smaller uncut chip thickness and a poor surface roughness.
|
|
| 6. |
- Liu, Yang, et al.
(författare)
-
Investigation on residual stress evolution in nickel-based alloy affected by multiple cutting operations
- 2021
-
Ingår i: Journal of Manufacturing Processes. - : Elsevier BV. - 1526-6125. ; 68, s. 818-833
-
Tidskriftsartikel (refereegranskat)abstract
- Machining-induced residual stresses in a part can significantly influence its performance and service time. It is common that the final surface of the part is produced after multiple cuts in the machining process. In multiple cuts, the previous cuts could often generate accumulated strain/stress and temperature on the part surface. These accumulated strain/stress and temperature will be brought to the subsequent cut or final cut and continuously affect the cutting forces, process temperatures, deformation zones, etc. in subsequent cut, and eventually affect the final residual stresses on the part. This paper reports on a joint experimental and numerical investigation to explore the influence of previous cuts on surface residual stresses with consideration of cutting parameters, cutting procedure, and tool geometries in multiple cut of Inconel 718 alloy. Coupled Eulerian and Lagrangian (CEL) formulation is used in a two/three-cuts numerical model. The loading cycles of the selected material nodes are characterised based on isotropic constitutive model (Johnson-Cook model) to analyse the underlying mechanism of residual stress evolution during cutting sequences. The results show that accumulated stress/strain induced by previous cuts lead to a more curled chip morphology, a slight decrease in cutting force and a slight increase in feed force in the subsequent cut. An increased magnitude and depth of compressive residual stresses in the finished workpiece are generated owing to the influence of previous cuts, and this is more obvious when the previous cut is implemented at a larger uncut chip thickness, using a more negative rake angle or a larger edge radius tool. The residual stress level might be controlled by optimizing the previous cuts to get the desired surface integrity.
|
|
| 7. |
- Liu, Yang, et al.
(författare)
-
Numerical and experimental investigation of tool geometry effect on residual stresses in orthogonal machining of Inconel 718
- 2021
-
Ingår i: Simulation Modelling Practice and Theory. - : Elsevier BV. - 1569-190X. ; 106
-
Tidskriftsartikel (refereegranskat)abstract
- Residual stress has become more important than ever with the increasing performance requirement of components especially for those applied in safety-critical areas. As the machining process is fundamentally correlated with the acquired component properties, it is essential to fully understand the formation mechanism of residual stresses in the cutting process and its influence on the performance of the component. This paper presents results based on numerical and experimental analysis on the effect of tool geometry on thermal-mechanical load and residual stresses in orthogonal machining Inconel718 alloy. The Coupled Eulerian-Lagrangian (CEL) method is used to simulate the effect of tool geometry on temperatures, forces, equivalent plastic strains, and residual stresses. The local normal/tangential stress is introduced to determine the degree of the tensile plastic deformation induced by the tool. It is observed that a negative rake angle and a sharp edge radius tool tend to generate more compressive stress on the machined surface than the ones generated with positive rake angle tools and/or lager edge radius. Besides, an increase in flank wear produces less magnitude of compressive stress in subsurface due to a decreased local normal stress caused by increased flank contact length.
|
|
| 8. |
- Liu, Yang, et al.
(författare)
-
Numerical contribution to segmented chip effect on residual stress distribution in orthogonal cutting of Inconel718
- 2020
-
Ingår i: International Journal of Advanced Manufacturing Technology. - : Springer Science and Business Media LLC. - 0268-3768 .- 1433-3015. ; 109, s. 993-1005
-
Tidskriftsartikel (refereegranskat)abstract
- Segmented chip morphology has a significant influence on distribution of residual stress and surface topography on machined surface in machining difficult-to-cut materials. In this paper, Coupled Eulerian and Lagrangian (CEL) model is employed to investigate the effect of segmented chip on surface integrity (residual stress distribution and surface topography) in orthogonal machining of Inconel718 with uncoated carbide inserts. A mesh sensitivity study of chip morphology and residual stress distribution is performed by developing the following three different grid resolutions: coarse (mesh size 35 μm), medium (10 μm) and fine (5 μm). Comparing with the experimental results, it is clear that the numerical model presents reasonable results, including the chip morphology, temperature distribution, cutting forces, residual stress profile and surface fluctuation period. As for the generated surface integrity, a waved surface and cyclic residual stress distribution are found with the segmented chip due to the periodical mechanical and thermal loadings acting on the machined surface. Furthermore, the formation of single chip segment is investigated in-depth to explain the residual stress distribution generation.
|
|
| 9. |
- Ren, Luyao, et al.
(författare)
-
Quartet DNA reference materials and datasets for comprehensively evaluating germline variant calling performance
- 2023
-
Ingår i: Genome Biology. - : BioMed Central (BMC). - 1465-6906 .- 1474-760X. ; 24:1
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Genomic DNA reference materials are widely recognized as essential for ensuring data quality in omics research. However, relying solely on reference datasets to evaluate the accuracy of variant calling results is incomplete, as they are limited to benchmark regions. Therefore, it is important to develop DNA reference materials that enable the assessment of variant detection performance across the entire genome.RESULTS: We established a DNA reference material suite from four immortalized cell lines derived from a family of parents and monozygotic twins. Comprehensive reference datasets of 4.2 million small variants and 15,000 structural variants were integrated and certified for evaluating the reliability of germline variant calls inside the benchmark regions. Importantly, the genetic built-in-truth of the Quartet family design enables estimation of the precision of variant calls outside the benchmark regions. Using the Quartet reference materials along with study samples, batch effects are objectively monitored and alleviated by training a machine learning model with the Quartet reference datasets to remove potential artifact calls. Moreover, the matched RNA and protein reference materials and datasets from the Quartet project enables cross-omics validation of variant calls from multiomics data.CONCLUSIONS: The Quartet DNA reference materials and reference datasets provide a unique resource for objectively assessing the quality of germline variant calls throughout the whole-genome regions and improving the reliability of large-scale genomic profiling.
|
|
| 10. |
- Sitbon, Olivier, et al.
(författare)
-
Selexipag for the Treatment of Pulmonary Arterial Hypertension
- 2015
-
Ingår i: New England Journal of Medicine. - 0028-4793 .- 1533-4406. ; 373:26
-
Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: In a phase 2 trial, selexipag, an oral selective IP prostacyclin-receptor agonist, was shown to be beneficial in the treatment of pulmonary arterial hypertension.METHODS: In this event-driven, phase 3, randomized, double-blind, placebo-controlled trial, we randomly assigned 1156 patients with pulmonary arterial hypertension to receive placebo or selexipag in individualized doses (maximum dose, 1600 μg twice daily). Patients were eligible for enrollment if they were not receiving treatment for pulmonary arterial hypertension or if they were receiving a stable dose of an endothelin-receptor antagonist, a phosphodiesterase type 5 inhibitor, or both. The primary end point was a composite of death from any cause or a complication related to pulmonary arterial hypertension up to the end of the treatment period (defined for each patient as 7 days after the date of the last intake of selexipag or placebo).RESULTS: A primary end-point event occurred in 397 patients--41.6% of those in the placebo group and 27.0% of those in the selexipag group (hazard ratio in the selexipag group as compared with the placebo group, 0.60; 99% confidence interval, 0.46 to 0.78; P<0.001). Disease progression and hospitalization accounted for 81.9% of the events. The effect of selexipag with respect to the primary end point was similar in the subgroup of patients who were not receiving treatment for the disease at baseline and in the subgroup of patients who were already receiving treatment at baseline (including those who were receiving a combination of two therapies). By the end of the study, 105 patients in the placebo group and 100 patients in the selexipag group had died from any cause. Overall, 7.1% of patients in the placebo group and 14.3% of patients in the selexipag group discontinued their assigned regimen prematurely because of adverse events. The most common adverse events in the selexipag group were consistent with the known side effects of prostacyclin, including headache, diarrhea, nausea, and jaw pain.CONCLUSIONS: Among patients with pulmonary arterial hypertension, the risk of the primary composite end point of death or a complication related to pulmonary arterial hypertension was significantly lower with selexipag than with placebo. There was no significant difference in mortality between the two study groups. (Funded by Actelion Pharmaceuticals; GRIPHON ClinicalTrials.gov number, NCT01106014.).
|
|
