A tutorial and spreadsheet for the validation and bottom-up uncertainty evaluation of quantifications performed by instrumental methods of analysis based on linear weighted calibrations is presented. The developed tool automatically assesses if calibrator values uncertainty is negligible given instrumental signal precision, assesses signal homoscedasticity by the Levene's test, guides the selection of weighting factors and evaluates the fitness of the regression model to define the calibration curve. The spreadsheet allows the use of the linear weighted regression model without the need for collecting many replicate signals of calibrators and sample by taking previously developed detailed models of signal precision variation in the calibration interval after adjustments to the daily precision conditions. This tool was successfully applied to the determination of the mass concentration of Cd, Pb, As, Hg, Co, V and Ni in a nasal spray by ICP-MS after samples dilution and acidification. The developed uncertainty models were checked through the analysis of nasal sprays after spiking with known analyte concentration levels. The metrological compatibility between estimated and reference analyte levels for 95% or 99% confidence level supports uncertainty model adequacy. The spiked samples were quantified from many replicate signals but uncertainty evaluation from duplicate calibrator and sample signals was assessed by randomly selecting calibrators and sample signals and by numerically defining a minimum acceptable success rate of the compatibility tests. The developed model was proven adequate to quantify the uncertainty of the studied measurements.

• MeSH
• Calibration MeSH
• Linear Models MeSH
• Uncertainty MeSH
• Nasal Sprays * MeSH
• Spectrum Analysis MeSH
• Publication type
• Journal Article MeSH

Stanovení FreeLite Chains (FLC) nachází v klinické praxi stále širší uplatnění. Stává se vedle elektroforézy bílkovin séra a imunofixační elektroforézy součástí diagnostiky, prognostiky a monitorování terapie u monoklonálních gamapatií. Vlastní stanovení FLC má však zatím četná metodická úskalí a je snaha o jejich postupné odstranění. Touto snahou byly také vedeny Česká myelomová skupina a Česká společnost klinické biochemie, když organizovaly studii, při které šest laboratoří center léčby mnohočetného myelomu v České republice současně vyšetřilo 12 vzorků sér od nemocných monoklonálními gamapatiemi na stanovení FLC. Z této studie vyplývají tyto závěry: jednotlivé laboratoře sjednotí ředění sér podle pokynů výrobce, index kappa/lambda je stanovitelný s menší přesností, proto je doporučeno používat ve větší míře hodnot koncentrací FLC. Do kontrolního cyklu SEKK (www.sekk.cz) Gamapatie je od počátku roku 2010 pravidelně zařazováno stanovení FLC.

Free monoclonal immunoglobulin light chains (FLC) quantification is recommended for diagnosis assessment and monitoring therapy for patients with monoclonal gammopathy. But there are numerous uncertainties regarding the detection, interpretation and FLC quantification. Our interlaboratory study showed that kappa/lambda ratio is strongly influenced by measurement errors and therefore we recommended the preferential use of FLC concentration values. Unified protocols are needed to minimise interlaboratory variability introduced by manual dilution or volume augmentation of clinical sample.

• MeSH
• Amyloidosis diagnosis   etiology   immunology   MeSH
• Financing, Organized MeSH
• Clinical Laboratory Techniques trends   utilization   MeSH
• Immunoglobulin Light Chains blood   MeSH
• Humans MeSH
• Multiple Myeloma diagnosis   etiology   immunology   MeSH
• Antibodies, Monoclonal therapeutic use   MeSH
• Paraproteinemias diagnosis   immunology   MeSH
• Paraproteins isolation & purification   MeSH
• Reproducibility of Results MeSH
• Quality Control MeSH
• Societies, Medical MeSH
• Statistics as Topic MeSH
• Validation Studies as Topic MeSH
• Check Tag
• Humans MeSH

Serial quantification of BCR-ABL1 mRNA is an important therapeutic indicator in chronic myeloid leukaemia, but there is a substantial variation in results reported by different laboratories. To improve comparability, an internationally accepted plasmid certified reference material (CRM) was developed according to ISO Guide 34:2009. Fragments of BCR-ABL1 (e14a2 mRNA fusion), BCR and GUSB transcripts were amplified and cloned into pUC18 to yield plasmid pIRMM0099. Six different linearised plasmid solutions were produced with the following copy number concentrations, assigned by digital PCR, and expanded uncertainties: 1.08±0.13 × 10(6), 1.08±0.11 × 10(5), 1.03±0.10 × 10(4), 1.02±0.09 × 10(3), 1.04±0.10 × 10(2) and 10.0±1.5 copies/μl. The certification of the material for the number of specific DNA fragments per plasmid, copy number concentration of the plasmid solutions and the assessment of inter-unit heterogeneity and stability were performed according to ISO Guide 35:2006. Two suitability studies performed by 63 BCR-ABL1 testing laboratories demonstrated that this set of 6 plasmid CRMs can help to standardise a number of measured transcripts of e14a2 BCR-ABL1 and three control genes (ABL1, BCR and GUSB). The set of six plasmid CRMs is distributed worldwide by the Institute for Reference Materials and Measurements (Belgium) and its authorised distributors (https://ec.europa.eu/jrc/en/reference-materials/catalogue/; CRM code ERM-AD623a-f).

• MeSH
• Fusion Proteins, bcr-abl genetics   metabolism   MeSH
• DNA MeSH
• Gene Dosage MeSH
• Calibration MeSH
• Cloning, Molecular MeSH
• Real-Time Polymerase Chain Reaction standards   MeSH
• Humans MeSH
• Membrane Transport Proteins genetics   MeSH
• RNA, Messenger metabolism   MeSH
• Plasmids genetics   MeSH
• Escherichia coli Proteins genetics   MeSH
• Proto-Oncogene Proteins c-bcr genetics   MeSH
• Reference Standards MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Three alternative procedures were employed for the isolation of polycyclic aromatic hydrocarbons (PAHs; 15 of 16 US EPA priority pollutants and benzo[e]pyrene), their methyl-derivatives and sulphur analogues from fish tissue: (1) Soxhlet extraction, (2) batch extraction enhanced by sonication, and (3) saponification of the sample followed by re-extraction of analytes into hexane. Soxhlet extraction using hexane-acetone (1:1, v/v) was the most efficient extraction technique, with analyte recoveries in the range 70-108%. Within optimization of the clean-up step, several types of gel permeation chromatography (GPC) systems were tested: two types of polystyrene divinylbenzene copolymer gels (PSDVB), both 'soft' gel type (Bio-Beads S-X3) and 'rigid' gels type (PL gel and Envirogel) in combination with various mobile phases were compared. Bio-Beads S-X3 and mobile phase chloroform were the most appropriate for purifying of crude extracts before the final determinative step. High-performance liquid chromatography with fluorimetric detection (HPLC/FLD) was used for identification and quantification of PAHs in purified fish extracts. The uncertainties of PAHs measurements were estimated by employing two alternative approaches. Both provided similar results: the expanded uncertainties obtained for individual PAHs by the 'top-down' approach were in the range 9-53%, their values resulting from application of the 'bottom-up' approach were in the range 16-52%.

• MeSH
• Chloroform MeSH
• Chromatography, Gel methods   MeSH
• Indicators and Reagents MeSH
• Carcinogens, Environmental analysis   MeSH
• Food Contamination analysis   MeSH
• Mutagens analysis   MeSH
• Noxae analysis   MeSH
• Perciformes MeSH
• Polycyclic Aromatic Hydrocarbons analysis   MeSH
• Polystyrenes MeSH
• Solvents MeSH
• Fish Products analysis   MeSH
• Sonication MeSH
• Chromatography, High Pressure Liquid methods   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH

PURPOSE: The aims of this work are (1) to explore deep learning (DL) architectures, spectroscopic input types, and learning designs toward optimal quantification in MR spectroscopy of simulated pathological spectra; and (2) to demonstrate accuracy and precision of DL predictions in view of inherent bias toward the training distribution. METHODS: Simulated 1D spectra and 2D spectrograms that mimic an extensive range of pathological in vivo conditions are used to train and test 24 different DL architectures. Active learning through altered training and testing data distributions is probed to optimize quantification performance. Ensembles of networks are explored to improve DL robustness and reduce the variance of estimates. A set of scores compares performances of DL predictions and traditional model fitting (MF). RESULTS: Ensembles of heterogeneous networks that combine 1D frequency-domain and 2D time-frequency domain spectrograms as input perform best. Dataset augmentation with active learning can improve performance, but gains are limited. MF is more accurate, although DL appears to be more precise at low SNR. However, this overall improved precision originates from a strong bias for cases with high uncertainty toward the dataset the network has been trained with, tending toward its average value. CONCLUSION: MF mostly performs better compared to the faster DL approach. Potential intrinsic biases on training sets are dangerous in a clinical context that requires the algorithm to be unbiased to outliers (i.e., pathological data). Active learning and ensemble of networks are good strategies to improve prediction performances. However, data quality (sufficient SNR) has proven as a bottleneck for adequate unbiased performance-like in the case of MF.

• MeSH
• Algorithms MeSH
• Deep Learning * MeSH
• Bias MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

PURPOSE: Computational models of microwave tissue ablation are widely used to guide the development of ablation devices, and are increasingly being used for the development of treatment planning and monitoring platforms. Knowledge of temperature-dependent dielectric properties of lung tissue is essential for accurate modeling of microwave ablation (MWA) of the lung. METHODS: We employed the open-ended coaxial probe method, coupled with a custom tissue heating apparatus, to measure dielectric properties of ex vivo porcine and bovine lung tissue at temperatures ranging between 31 and 150  ∘ C, over the frequency range 500 MHz to 6 GHz. Furthermore, we employed numerical optimization techniques to provide parametric models for characterizing the broadband temperature-dependent dielectric properties of tissue, and their variability across tissue samples, suitable for use in computational models of microwave tissue ablation. RESULTS: Rapid decreases in both relative permittivity and effective conductivity were observed in the temperature range from 94 to 108  ∘ C. Over the measured frequency range, both relative permittivity and effective conductivity were suitably modeled by piecewise linear functions [root mean square error (RMSE) = 1.0952 for permittivity and 0.0650 S/m for conductivity]. Detailed characterization of the variability in lung tissue properties was provided to enable uncertainty quantification of models of MWA. CONCLUSIONS: The reported dielectric properties of lung tissue, and parametric models which also capture their distribution, will aid the development of computational models of microwave lung ablation.

• MeSH
• Ablation Techniques * MeSH
• Models, Biological * MeSH
• Electric Impedance MeSH
• Microwaves MeSH
• Uncertainty MeSH
• Lung cytology   radiation effects   MeSH
• Swine MeSH
• Cattle MeSH
• Temperature * MeSH
• Animals MeSH
• Check Tag
• Cattle MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

• MeSH
• Environmental Health statistics & numerical data   MeSH
• Risk Assessment MeSH
• Environmental Pollutants adverse effects   toxicity   MeSH
• Toxicity Tests MeSH
• Environmental Exposure MeSH

• Conspectus
• Veřejné zdraví a hygiena
• NML Fields
• environmentální vědy
• chemie, klinická chemie
• NML Publication type
• publikace WHO

• MeSH
• Chemical Actions and Uses MeSH
• Risk Assessment MeSH
• Hazardous Substances administration & dosage   MeSH
• Environmental Exposure MeSH

• Conspectus
• Životní prostředí a jeho ochrana
• NML Fields
• chemie, klinická chemie
• environmentální vědy
• NML Publication type
• publikace WHO

• MeSH
• Dermatitis, Allergic Contact MeSH
• Risk Assessment MeSH
• Hazardous Substances toxicity   MeSH
• Radiodermatitis MeSH

• Conspectus
• Patologie. Klinická medicína
• NML Fields
• dermatovenerologie
• toxikologie
• NML Publication type
• publikace WHO

We have examined the imprecision in the estimation of PCR efficiency by means of standard curves based on strategic experimental design with large number of technical replicates. In particular, how robust this estimation is in terms of a commonly varying factors: the instrument used, the number of technical replicates performed and the effect of the volume transferred throughout the dilution series. We used six different qPCR instruments, we performed 1-16 qPCR replicates per concentration and we tested 2-10 μl volume of analyte transferred, respectively. We find that the estimated PCR efficiency varies significantly across different instruments. Using a Monte Carlo approach, we find the uncertainty in the PCR efficiency estimation may be as large as 42.5% (95% CI) if standard curve with only one qPCR replicate is used in 16 different plates. Based on our investigation we propose recommendations for the precise estimation of PCR efficiency: (1) one robust standard curve with at least 3-4 qPCR replicates at each concentration shall be generated, (2) the efficiency is instrument dependent, but reproducibly stable on one platform, and (3) using a larger volume when constructing serial dilution series reduces sampling error and enables calibration across a wider dynamic range.

• Publication type
• Journal Article MeSH