Semi-quantitative Results Research Articles

Flow behavior and strain rate sensitivity assessment of γ and γʹ phases in Co–Al–W-based superalloy using experimental and computational approaches

The recently developed precipitation-hardened cobalt-based superalloys can be considered as a new category of elevated temperature materials. An attempt was made in this research to estimate the elastoplastic characteristics of γ and γʹ phases embedded in Co–Al–W superalloy by using nanoindentation data and constitutive equations. The elastic modulus and hardness were initially utilized to determine the yield stress, which will be subsequently used to characterize the flow behavior in the plastic region. The proposed computational model has been verified with quantitative and semi-quantitative results, calculated by finite element analysis (FEA). It was perceived that both γ and γʹ phases in the investigated superalloy exhibited the ductile load-bearing response as γʹ strengthening phase displayed higher yield stress compared to the γ matrix. The work hardening rate and work hardening exponent were also identified to be higher for γʹ rather than γ phase due to the presence of an anti-phase boundary, which consequently promotes the strain hardening mechanism. The load–displacement curves in both phases illustrated pop-ins which marks the transition from elastic to plastic deformation. Finally, the elastic and plastic data of both constituent phases were incorporated into the FEA paradigm to simulate the indentation projected area. Based on the indentation morphology, the FEA simulation results were further validated with experimental nano-mechanical data. The results revealed that the load–displacement curve acquired from the numerical simulation of the Berkovich indenter has an excellent agreement with the obtained nanoindentation test. The primary purpose of this research is to recognize the correlation between the strain-rate sensitivity of the hardness and the flow stress using FEA results integrated with the analytical modeling.

Diagnostic Performance of Xpert MTB/RIF Ultra Compared with Predecessor Test, Xpert MTB/RIF, in a Low TB Incidence Setting: a Retrospective Service Evaluation.

ABSTRACTThe aim of this study was to evaluate the performance of Xpert MTB/RIF Ultra (Ultra) compared with its predecessor, Xpert MTB/RIF (Xpert), in the diagnosis of tuberculosis (TB) in a low TB incidence country. Retrospective analysis was performed on 689 clinical samples received between 2015 and 2018, on which Xpert was performed, and on 715 samples, received between 2018 and 2020, on which Ultra was performed. Samples were pulmonary (n = 830) and extrapulmonary (n = 574) in nature, and a total of 264 were culture positive for Mycobacterium tuberculosis complex (MTBC). The diagnostic performance of both assays was analyzed using culture as the reference standard. The sensitivity of Ultra for culture positive (smear positive and smear negative) MTBC samples, was 93.2% (110/118) compared with 82.2% (120/146) for Xpert (P = 0.0078). In smear negative-culture positive samples, Ultra had a sensitivity of 74.2% (23/31) versus 36.11% (13/36) for Xpert (P = 0.0018). Specificity of both assays was comparable at 94.8% (566/597) for Ultra and 95.8% (520/543) for Xpert (P = 0.4475). The sensitivity of Ultra and Xpert assays among exclusively pulmonary samples was 95.3% (82/86) and 90.3% (84/93), respectively (P = 0.1955), and 87.5% (28/32) and 67.9% (36/53), respectively, among extrapulmonary samples (P = 0.0426). Ultra showed improved performance compared with Xpert in a low TB incidence setting, particularly in smear negative and extrapulmonary MTBC disease. The specificity of Ultra was lower than Xpert, however, this was not statistically significant.IMPORTANCE The study demonstrates the improved sensitivity of the Ultra compared with the Xpert, particularly in smear negative TB disease, for both pulmonary and extrapulmonary samples in a low TB incidence setting. Cycle threshold (Ct) value for both assays was found to positively correlate with time to TB culture positivity, suggesting that Ct and semiquantitative results could be used as indicators of sample MTBC bacillary burden, and thus, perhaps, of transmission potential. This may have implications for the designation of patient isolation precautions.

Analysis of Elements in Snack Foods: A Closer Look at Pepperoni, Rice Noodles, Frozen Dinners, and Pizza

A set of food samples with different percentage compositions of fats, proteins, and carbohydrates were digested in a single microwave digestion batch and analyzed to determine elemental concentrations using inductively coupled plasma–mass spectrometry (ICP-MS) and ICP-optical emission spectroscopy (OES) using the U.S. Food and Drug Administration (FDA) Elemental Analysis Manual (EAM) 4.7 (for ICP-MS) and FDA EAM 4.4 (for ICP-OES). Built-in software tools were used to streamline the analytical workflow, which is especially useful for new or less experienced users running these methods. Processed foods often contain high concentrations of some elements and trace amounts of others, so a wide analytical range is required. The ICP-MS method used to analyze the varied samples in this work was new. To ensure optimum setup of the ICP-MS method, semi-quantitative results were used to assess the solid content of the food digests before the sample preparation dilution was finalized. The same food samples were analyzed using ICP-MS and ICP-OES and the two methods were compared.

Evaluation of a Cryptococcus capsular polysaccharide detection FungiXpert LFA (lateral flow assay) for the rapid diagnosis of Cryptococcosis.

The FungiXpert LFA was developed to diagnose fungal meningitis caused by Cryptococcus yeasts, by using serum or cerebrospinal fluid. It was compared to an existing lateral flow assay (LFA). The FungiXpert LFA performed well in qualitative and semi-quantitative tests.

Rapid segmentation and sensitive analysis of CRP with paper-based microfluidic device using machine learning

Microfluidic paper-based analytical devices (μPADs) have been widely used in point-of-care testing owing to their simple operation, low volume of the sample required, and the lack of the need for an external force. To obtain accurate semi-quantitative or quantitative results, μPADs need to respond to the challenges posed by differences in reaction conditions. In this paper, multi-layer μPADs are fabricated by the imprinting method for the colorimetric detection of C-reactive protein (CRP). Different lighting conditions and shooting angles of scenes are simulated in image acquisition, and the detection-related performance of μPADs is improved by using a machine learning algorithm. The You Only Look Once (YOLO) model is used to identify the areas of reaction in μPADs. This model can observe an image only once to predict the objects present in it and their locations. The YOLO model trained in this study was able to identify all the reaction areas quickly without incurring any error. These reaction areas were categorized by classification algorithms to determine the risk level of CRP concentration. Multi-layer perceptron, convolutional neural network, and residual network algorithms were used for the classification tasks, where the latter yielded the highest accuracy of 96%. It has a promising application prospect in fast recognition and analysis of μPADs.

Revising the performance of the Landau–Zener surface hopping on some typical one-dimensional nonadiabatic models

We implement the Landau–Zener surface hopping (LZSH) method to study the nonadiabatic dynamics of some typical one-dimensional (1D) models, including well-tested Tully's models. For models like the original linear avoided crossing model used to derive the LZ formula, the LZSH method can provide very accurate results compared with full quantum calculations and fewest-switches surface hopping (FSSH) simulations. For the Tully's single avoided crossing model, the LZSH method can only present us semi-quantitative results. And for Tully's dual avoided crossing model, the LZSH fails to capture the quantum interference effect. Furthermore, the LZSH shows a better performance for a three-state model compared with FSSH due to no coherence when the nuclear momentum is small. The simulation results show us the performance of LZSH method depends on the shape of adiabatic potential energy surface, which deepens our understanding of nonadiabatic dynamics for some typical 1D models and provides solid basics for further developing new methods based on the LZ formula.

Immunohistochemical staining of immunoglobulin G in healthy equine, canine, and feline corneas.

ObjectiveEstablishing an immunohistochemical approach for semi‐quantitative assessment of the presence of immunoglobulin G (IgG) in equine, canine, and feline corneas.ProceduresHealthy corneas of horses, dogs, and cats, euthanized because of a fatal disease or an unrecoverable trauma unrelated to and without a history of ophthalmic disease were formalin‐fixed, paraffin‐embedded, and determined to be pathomorphologically healthy by light microscopy. Automated immunohistochemistry was performed using primary antibodies against IgG, biotin‐conjugated secondary antibodies, and streptavidin‐peroxidase, as well as diaminobenzidine for visualization. After counterstaining with hematoxylin, epithelium, stroma, Descemet´s membrane (DM), and endothelium were semi‐quantitatively scored for the presence of IgG on a 4‐grade scale (0 = no, 1 = faint, 2 = medium, 3 = strong staining) by light microscopy.ResultsCorneal specimens of 20 horses (40 eyes) with a median age of 15.5 years (range 2–31 years), 12 dogs (21 eyes) with a median age of 10.0 years (range 4–16), and 13 cats (24 eyes) with a median age of 10.0 years (range 2–18) were included in the study. Different sexes and breeds were represented. In all corneas (100%), significant medium signal intensity in the stroma was observed. Variable immunosignal was obtained in epithelium, DM, and endothelium.ConclusionThis method reproducibly allows for the detection of IgG in healthy equine, canine, and feline corneas, particularly stroma. Semi‐quantitative results evidence medium presence of IgG in the corneal stroma. Further research is needed to evaluate IgG presence in diseased corneas.

A method comparison of three immunoassays for detection of neutralizing antibodies against SARS-CoV-2 receptor-binding domain in individuals with adenovirus type-5-vectored COVID-19 vaccination.

ObjectiveDetecting neutralizing antibodies targeting receptor‐binding domain (RBD) is important for the assessment of humoral protection and vaccine efficacy after vaccination. We compared the performance of three surrogate immunoassays for detection of neutralizing antibodies targeting RBD.MethodsWe analyzed 115 serum samples obtained from individuals with Ad5‐vectored COVID‐19 vaccination using two competitive enzyme‐linked immunosorbent assays (Wantai BioPharm and Synthgene Medical Technology) and one competitive chemiluminescence assay (YHLO Biotech). Performance evaluation and methodology comparison were performed according to the Clinical and Laboratory Standards Institute related guidelines.ResultsThe precision met the manufacturers’ statements. The linear range of the WANTAI was 0.0625–0.545 U/ml and the YHLO was 0.260–242.4 U/ml. The WANTAI’s limit of blank (LoB) and limit of detection (LoD) were 0.03 and 0.06 U/ml, respectively. The YHLO’s LoB and LoD were 0.048 and 0.211 U/ml, respectively. The correlations of semi‐quantitative results of Synthgene with quantitative results of YHLO (ρ = 0.566) and WANTAI (ρ = 0.512) were medium. For YHLO and WANTAI, there was a good agreement (0.62) and a strong correlation (ρ = 0.931). Passing–Bablok analysis and Bland‐Altman plot showed a positive bias (112.3%) of the YHLO compared to the WANTAI. The exclusion of samples >50 U/ml did not decrease bias.ConclusionThese findings contribute to a deeper understanding of surrogate viral neutralization assays and provide useful data for future comparison studies.

Abstract P1-06-07: Enhanced differentiation of clinical behavior by breast carcinomas utilizing quantification of estrogen and progestin receptor proteins and gene expression subsets

Abstract Background: IHC analyses of clinical biomarkers for breast cancer management yield semi-quantitative results potentially complicated by variation in methods and interpretation. We are mining a comprehensive database established over four decades that contains deidentified results of quantified protein and RNA biomarkers with clinical outcomes to detect candidates for companion diagnostics and molecular targets. Assays were performed in a Reference Laboratory holding both CLIA and Commonwealth of Kentucky licenses. Methods: This retrospective investigation utilized ER/PR protein levels, expressed as fmol/mg cytosol protein, that were quantified for each of 2339 carcinoma biopsies with FDA-approved kits and cut-off values by radio-ligand binding assay (LBA, NEN/DuPont) and/or enzyme immunoassay (EIA, Abbott Labs). Of these, 454 specimens had both LBA and EIA results recorded. Biomarker results and de-identified clinical outcomes were examined using REMARK criteria. Gene expression, quantified biomarkers, features of primary breast cancers and clinical patient outcomes were analyzed by univariable and multivariable Cox regressions, Fisher’s Exact Test, Kaplan Meier plots and with R software v4.0.0. Microarray results of ~22,000 genes were obtained from RNA of LCM-procured breast carcinoma cells of 247 primary breast biopsies and expression levels of 85 cancer-associated genes were determined by RTqPCR of 278 specimens after RNA extraction and purification with established procedures. Results: Quantified levels of both ER and PR protein were expressed to a greater extent in primary carcinomas from post-menopausal patients compared to cancers from premenopausal women regardless of assay type (p = 0.005). ESR1 expression by microarray was only greater in LCM-procured carcinoma cells of post-menopausal patients while both ESR1 and PGR relative expression levels measured by qPCR were greater in tissue biopsies of post-menopausal women (p = 0.05). Box plots of untransformed EIA/LBA results of either ER (n=454) or PR (n=452) using specimens on which both quantifying assays were performed revealed a significant positive ER bias by EIA (median ratio=1.54) compared to that of PR (median ration=1.13). Transformed data of ratios confirmed this observation. Kaplan Meier plots of patient outcomes with ER or PR or both assessed by each assay type suggested LBA was a slightly better predictor. Microarray analyses of LCM-procured cells revealed 2994 genes distinguishing ER+ from ER- breast cancers of which ~200 genes were identified that differentiated 4 distinct molecular subtypes (2 ER+ and 2 ER-) exhibiting different outcomes upon Kaplan-Meier analyses. Expression levels of the 85 genes (including ERBB2), validated by qPCR with RNA from biopsy tissue sections, were evaluated relative to quantified levels of either ER or PR protein or both in primary breast cancer biopsies. Collectively, use of quantified ER/PR protein or ESR1/PGR expression with clinical outcomes enhanced identification of small molecular signatures and drug development candidates. Conclusions: Measurements of ER, PR and HER2 proteins with tests that quantify their levels as well as by microarray and qPCR assessment of ESR1, PGR and ERBB2 relative gene expression were correlated with prediction of clinical outcomes of breast carcinomas exhibiting various features (e.g., pathology classification, TNBC). Results reinforced importance of assessing levels of the clinically relevant biomarkers in a quantified fashion to enhance their use in breast cancer management and prediction of risk of recurrence. These investigations also revealed numerous over-expressed genes in carcinomas with poor clinical outcomes suggesting candidates for development of novel therapeutics. Citation Format: James L Wittliff, Michael W. Daniels. Enhanced differentiation of clinical behavior by breast carcinomas utilizing quantification of estrogen and progestin receptor proteins and gene expression subsets [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P1-06-07.

Optimization of image reconstruction method of cerebral blood flow perfusion imaging with digital CZT SPECT.

The purpose of this study is to evaluate the effects of filtered back projection (FBP), ordered subset expectation maximisation (OSEM), and different filters on cadmium zinc telluride single-photon emission computed tomography [CZT single-photon emission computed tomography (SPECT)] cerebral blood perfusion image quality to optimise the image reconstruction method. Under routine clinical conditions, tomographic imaging was performed on the phantom and patients. Image processing included image reconstruction using FBP and OSEM, and the filtering method used Butterworth (Bw) and Gaussian (Gs) filters. Visual and semi-quantitative parameters [integral uniformity, root mean square (RMS) noise and contrast and contrast-to-noise ratio (CNR)] were used to evaluate image quality to optimise image reconstruction parameters. One-way and two-way analysis of variance were used to process phantom and clinical data. In the tomographic images of the phantom, the semi-quantitative analysis showed that the integral uniformity of FBP+Bw was better than that of OSEM+Bw and OSEM+Gs (P < 0.05), and that the RMS noise of FBP+Bw was lower than that of OSEM+Bw and OSEM+Gs (P < 0.001). The contrast of FBP+Bw and OSEM+Bw in the cold area diameter ≥2 cm group was higher than that of OSEM+Gs (P < 0.001), whereas the CNR of FBP+Bw was higher than that of OSEM+Bw and OSEM+Gs (P < 0.001); the contrast of OSEM+Bw cold area diameter <2 cm was higher than that of FBP+Bw (P < 0.01). The semi-quantitative analysis results of the clinical images were consistent with the phantom's. In CZT SPECT cerebral blood flow perfusion imaging, it is suggested that the image postprocessing method of FBP+Bw (fc = 0.40; n = 10) should be used routinely in clinical application, and if there are uncertain small lesions in the processed image, it is suggested to use the reconstruction method of OSEM+Bw (EM-equivalent iterations = 60; fc = 0.45; n = 10) instead.

Effect of not vortexing nasopharyngeal and throat swabs on SARS-CoV-2 nucleic acid detection: A pilot study

The processing of swabs for respiratory virus detection involves vortexing while still in the viral transport medium (VTM). The effect of not vortexing swabs prior to analysis has not been studied extensively for SARS-CoV-2 detection, and presents an opportunity to improve pre-analytic laboratory workflow. We aimed to assess the impact of not vortexing nasopharyngeal/throat swabs submitted in VTM for SARS-CoV-2 testing. To assess the impact of not vortexing swabs, 277 swab samples were tested for SARS-CoV-2 RNA in paired vortexed and non-vortexed aliquots using eight routine nucleic acid amplification assays. We compared the qualitative (positive/negative) and semi-quantitative (cycle threshold, Ct) results. Following discordant analysis, all but one non-vortexed sample had the same qualitative result as the vortexed sample. 27.4 % of samples were SARS-CoV-2 positive. Comparison of Ct values revealed an apparent reduction in human cellular nucleic acid in the non-vortexed samples (mean Ct values of 24.0 and 26.5 for vortexed and non-vortexed samples, respectively, p < 0.0001) and increased Ct values for non-vortexed samples using a laboratory-developed SARS-CoV-2 assay (mean Ct values of 4.1 and 4.2 for vortexed and non-vortexed samples, respectively; p < 0.0001), but this was not observed for a more automated commercial SARS-CoV-2 assay (mean Ct values of 15.2 for both vortexed and non-vortexed samples, respectively; p = 0.68). While vortexing swabs appears to improve the recovery of cellular material, it does not have an appreciable impact on the qualitative sensitivity of SARS-CoV-2 nucleic acid tests, which may support omission of this step and simplification of front-end sample processing.

Using equilibrium thermodynamics to study thermochemical processes in arc discharges used in atomic emission spectral analysis (review)

Until the beginning of the 21st century, the arc discharge was the main source of excitation of the atomic spectra of elements during routine atomic emission spectral analysis. The wide application of this spectral source at the same time aroused great interest of analysts in the study of complex thermochemical processes occurring at its electrodes and in the discharge plasma. This was due to the need to improve the metrological and informational characteristics of the analysis. Equilibrium thermodynamics has often been used to carry out such studies. It is shown that three levels of the thermodynamic approach can be distinguished here, differing in the complexity of the applied apparatus of thermodynamics and the information content of the results obtained. Numerous examples of the application of these levels of approach to the study of thermochemical processes occurring in the electrode or in the discharge plasma are given. The main advantages and disadvantages of the approaches used, which provide only qualitative information about the ongoing processes, are noted. The main disadvantage is the impossibility of predicting the composition of the arc plasma from the initial composition of the condensed sample. The most informative is the generalized thermodynamic model of thermochemical processes in a DC arc discharge from the sample surface and from the electrode crater. The model is based on the division of a non-stationary non-equilibrium system of an arc discharge into successive quasi-stationary subsystems. They correspond to the stage of sample evaporation and thermochemical transformation of its components in the discharge plasma. According to the model, the total equilibrium composition of the evaporated sample is initially calculated. This determines the initial composition of the arc plasma. Then, the total equilibrium composition of the arc plasma is calculated and the transition to the intensities of the spectral lines of the analytes is performed. The correctness of the generalized model, which gives semi-quantitative results, is confirmed by comparison with experimental data.

Establishment of Doxorubicin Resistant Stem-like Cell line from MCF-7 Cells

Background Conventional therapy of Breast cancer (BCa) is associated with numerous challenges, relapse being the topmost. It has been shown that disease relapse occurs due to the existence of cancer stem cells (CSCs), residing within the tumor. Objectives To develop a cancer stem cell line model, exploiting the property of stem cells to resist treatment with anti-cancer therapeutic agents and to develop novel therapeutic interventions targeting the elimination of CSCs. Methods Single-cell clones (SCC) from MCF-7 cells were established in 96-well plates using a culture medium containing 5% FBS. Enrichment of cancer stem cells was carried out from established single cell clones which were initially cultured in a serum-deprived nutrient medium for six weeks followed by Doxorubicin treatment. Doxorubicin-resistant clones were established and evaluated for their growth and sphere-forming abilities. Further, these clones were characterized based on the presence of stem-cell markers using a semi-quantitative reverse transcription-polymerase chain reaction and results were compared with the parental MCF-7 cell line. Results In a complete medium, these spheres differentiated and started growing as a monolayer with differential expression levels of genes involved in stemness and cellular differentiation. When these spheres were sub-cultured again in stem-cell medium and detached to give rise to single cells, these clones retained sphere-forming ability. Doxorubicin-resistant clones showed a tendency to grow in spheres in a stem-cell medium with serum-deprived culture conditions. Conclusion Our results provide undeviating evidence of the successful establishment of clones of MCF-7 cell line exhibiting stem cell-like properties. These cell lines have been reprogrammed as stem cell-like models and can be used to delineate recurrent breast cancer attributed to CSCs.

Physicochemical characterization of short and long-term aged asphalt mixtures for low-temperature performance

Current pavement engineering shift towards Balanced Mix Design warrants performance testing of asphalt mixtures at low temperatures. The Bending Beam Rheometer (BBR) is an effective tool to characterize asphalt mixture’s low-temperature properties based on AASHTO TP125. This study utilized the BBR for physical characterization of 32 different asphalt mixtures composed of a PG 64–28 binder, two Job-Mix Formulae (JMF), four different RAP contents, and four different aging methods. Chemically extracted binder from the aged mixtures was then analyzed using the absorbance-based Fourier-Transform Infrared Spectroscopy (FT-IR) to identify the critical functional groups. The FT-IR analysis's semi-quantitative results were validated using the Energy Dispersive X-Ray Analyzer on the Scanning Electron Microscope (SEM-EDX). The reduction of carbon and sulfur concentration due to oxidation was correlated to the loss of cohesion between asphalt mastic and aggregates. Furthermore, microstructural damage was found in the aged specimens at a 200x magnification. Overall, this study provides a comprehensive and feasible method to perform physicochemical characterization of asphalt mixtures.

RLEP LAMP for the laboratory confirmation of leprosy: towards a point-of-care test

BackgroundNucleic acid-based amplification tests (NAAT), above all (q)PCR, have been applied for the detection of Mycobacterium leprae in leprosy cases and household contacts with subclinical infection. However, their application in the field poses a range of technical challenges. Loop-mediated isothermal amplification (LAMP), as a promising point-of-care NAAT does not require sophisticated laboratory equipment, is easy to perform, and is applicable for decentralized diagnosis at the primary health care level. Among a range of gene targets, the M. leprae specific repetitive element RLEP is regarded as highly sensitive and specific for diagnostic applications. MethodsOur group developed and validated a dry-reagent-based (DRB) RLEP LAMP, provided product specifications for customization of a ready-to-use kit (intended for commercial production) and compared it against the in-house prototype. The assays were optimized for application on a Genie® III portable fluorometer. For technical validation, 40 “must not detect RLEP” samples derived from RLEP qPCR negative exposed and non-exposed individuals, as well as from patients with other conditions and a set of closely related mycobacterial cultures, were tested together with 25 “must detect RLEP” samples derived from qPCR confirmed leprosy patients. For clinical validation, 150 RLEP qPCR tested samples were analyzed, consisting of the following categories: high-positive samples of multibacillary (MB) leprosy patients (> 10.000 bacilli/extract), medium-positive samples of MB leprosy patients (1.001–10.000 bacilli/extract), low-positive samples of MB leprosy patients (1–1.000 bacilli/extract), endemic controls and healthy non-exposed controls; each n = 30. ResultsTechnical validation: both LAMP formats had a limit of detection of 1.000 RLEP copies, i.e. 43–27 bacilli, a sensitivity of 92% (in-house protocol)/100% (ready-to-use protocol) and a specificity of 100%. Reagents were stable for at least 1 year at 22 °C. Clinical validation: Both formats showed a negativity rate of 100% and a positivity rate of 100% for high-positive samples and 93–100% for medium positive samples, together with a positive predictive value of 100% and semi-quantitative results. The positivity rate for low-positive samples was 77% (in-house protocol)/43% (ready-to-use protocol) and differed significantly between both formats. ConclusionsThe ready-to-use RLEP DRB LAMP assay constitutes an ASSURED test ready for field-based evaluation trials aiming for routine diagnosis of leprosy at the primary health care level.

The first report on the sortase-mediated display of bioactive protein A from Staphylococcus aureus (SpA) on the surface of the vegetative form of Bacillus subtilis

Protein A (SpA) is one of the most important Staphylococcus aureus cell wall proteins. It includes five immunoglobulin (Ig)-binding domains which can bind to immune complexes through the Fc region of immunoglobulins. The binding of SpA to the polymeric supports can be used to prepare affinity chromatography resins, which are useful for immunoprecipitation (IP) of antibodies. Protein A is also used to purify many anti-cancer antibodies. In this study, SpA was displayed on the surface of Bacillus subtilis cells using a sortase-mediated system to display the target protein to the B. subtilis cell wall. A series of plasmids consisting of cassettes for cell wall-directed protein A as well as negative controls were constructed and transformed into B. subtilis WASD (wprA sigD) cells. SDS-PAGE, western blot, flow cytometry, functional IgG purification assay, and a modified ELISA assay were used to confirm the surface display of SpA and evaluate its function. Semi-quantitative ELISA results showed that the binding capacity of lyophilized Bs-SpA is 100 μg IgG from rabbit serum per 1 mg of cells under optimal experimental conditions. Low production costs, optimal performance, and the use of a harmless strain compared to a similar commercial product predict the possible use of SpA immobilization technology in the future.

WebMaBoSS: A Web Interface for Simulating Boolean Models Stochastically.

WebMaBoSS is an easy-to-use web interface for conversion, storage, simulation and analysis of Boolean models that allows to get insight from these models without any specific knowledge of modeling or coding. It relies on an existing software, MaBoSS, which simulates Boolean models using a stochastic approach: it applies continuous time Markov processes over the Boolean network. It was initially built to fill the gap between Boolean and continuous formalisms, i.e., providing semi-quantitative results using a simple representation with a minimum number of parameters to fit. The goal of WebMaBoSS is to simplify the use and the analysis of Boolean models coping with two main issues: 1) the simulation of Boolean models of intracellular processes with MaBoSS, or any modeling tool, may appear as non-intuitive for non-experts; 2) the simulation of already-published models available in current model databases (e.g., Cell Collective, BioModels) may require some extra steps to ensure compatibility with modeling tools such as MaBoSS. With WebMaBoSS, new models can be created or imported directly from existing databases. They can then be simulated, modified and stored in personal folders. Model simulations are performed easily, results visualized interactively, and figures can be exported in a preferred format. Extensive model analyses such as mutant screening or parameter sensitivity can also be performed. For all these tasks, results are stored and can be subsequently filtered to look for specific outputs. This web interface can be accessed at the address: https://maboss.curie.fr/webmaboss/ and deployed locally using docker. This application is open-source under LGPL license, and available at https://github.com/sysbio-curie/WebMaBoSS.

The Effect of Prickly Ash (Zanthoxylum bungeanum Maxim) on the Taste Perception of Stewed Sheep Tail Fat by LC-QTOF-MS/MS and a Chemometrics Analysis.

This work aims to explore the contribution of prickly ash (Zanthoxylum bungeanum Maxim) on the taste perception of stewed sheep tail fat. Liquid chromatography-tandem quadrupole time of flight mass spectrometry (LC-QTOF-MS) was applied to analyze the taste-related compounds. A total of 99 compounds in different sheep tail fat samples were identified. The semi-quantitative results showed that there were differences between the samples. The partial least squares discriminant analysis (PLS-DA) model without overfitting was used to investigate the effect of prickly ash. Eleven marker compounds were predicted with a variable importance for projection > 1, fold change > 2 and p < 0.05. An additional experiment showed that guanosine 5′-monophosphate, malic acid, inosine and adenosine 5′-monophosphate could improve the umami and saltiness taste of stewed sheep tail fat.

Total oxidisable precursor assay towards selective detection of PFAS in AFFF

Total oxidisable precursor assay (TOP assay) can degrade and convert “unknown” per- and polyfluoroalkyl substances (PFAS) to detectable PFAS. However, the detailed degradation pathway is still not known, particularly when the TOP assay is applied to analyse complex samples such as aqueous film-forming foam (AFFF). To gain insights into the pathway and the effectiveness of the TOP assay, several “known” compounds are first tested as controls, including sodium dodecyl benzene sulphate (SDBS), perfluorooctanoic acid (PFOA), perfluorooctanesulfonic acid (PFOS). Secondly, the test is expanded to several PFAS precursors such as 6:2 fluorotelomer sulfonate (6:2 FTS), 8:2 fluorotelomer sulfonate (8:2 FTS), and a cationic surfactant N-ethyl-N-(2-hydroxyethyl) perfluorooctyl sulfonamide (EtFOSE). Thirdly, the TOP assay is used to test “unknown” PFAS samples that have been previously used as AFFF in Australia. The degradation products are monitored, to compare the mass balance and propose the degradation pathway. While HPLC-MS/MS is typically employed to detect the individual TOP assay products, most of which are perfluoroalkyl carboxylic acids (PFCA), an app-based smartphone sensor can also provide semi-quantitative results as a sum. Overall, the results indicate the effectiveness of the TOP assay to assess the presence of PFAS precursors in the AFFF samples, with some variations in the end products. Recommendations for enhancement of the TOP assay are also provided.

Need for caution when interpreting Xpert® MTB/RIF results for rifampin resistance among children

BACKGROUND: Recommended by the World Health Organization as an initial diagnostic test for TB in children, Xpert® MTB/RIF is widely implemented in many countries, including Kenya.METHODS: Three hundred HIV-positive and negative children (<5 years) were enrolled in Kisumu County, Kenya, from October 2013 to August 2015. Multiple specimen types were collected from each child and tested using Xpert, liquid culture, and phenotypic drug susceptibility testing (DST). Samples positive for rifampin (RIF) resistance on Xpert were tested using line-probe assay and sequencing.RESULTS: Of 32 children with bacteriologically confirmed TB, 27 had positive Xpert results. Of these, 3/27 (11%, 95% CI 4-28) had RIF resistance detected on Xpert, but not by phenotypic DST, line-probe assay, or sequencing. For these three children, five Xpert tests showed RIF resistance; all five tests had semi-quantitative "very low" results and delay or absence of probe D signal, whereas no Xpert results with higher semi-quantitative results showed RIF resistance. All three children responded well to standard TB treatment.CONCLUSIONS: False RIF resistance may be detected in pediatric specimens. Further study is needed to determine if false RIF resistance is associated with low bacterial load.