Standard Calibration Research Articles

Fraunhofer line-based wavelength-calibration method without calibration targets for planetary lander instruments

High-accuracy wavelength calibration is critical for qualitative and quantitative spectroscopic measurements. Many spectrometers employed in planetary-exploration missions have onboard calibration sources, including standard lamps and calibration targets. However, such calibration sources are not always available because planetary missions, particularly landing missions, usually have limitations in size and mass. Thus, a wavelength calibration method without requiring hardware addition can be highly beneficial. In this study, we demonstrate a method for wavelength calibration using solar Fraunhofer lines observed in the reflectance spectra of planetary surfaces. Using a Raman spectrometer prototype developed for a Phobos rover, we measured the spectrum of the sunlight reflected from a spectral standard, manufactured to provide similar reflectance spectra to the surface of Phobos. We identified 35 Fraunhofer absorption lines in the wavelength range between 530 and 700 nm and utilized these features for the wavelength calibration of the spectrometer. This approach using Fraunhofer lines achieved good results (better than +0.04/−0.06 nm), comparable to the results achieved using a conventional Ne lamp. The wavelength accuracy corresponds to a wavenumber accuracy better than ±1.5 cm−1 in the 0–4000 cm−1 Raman shift (Stokes shift) range with a 532 nm excitation laser. This result enabled the estimation of the magnesium number (Mg#) of olivine, achieving a value more precise than 1.5% based on the Raman peak positions. In addition, we examined the number of solar Fraunhofer lines detectable at different wavelength resolutions by binning the solar spectrum acquired in this study. We found that more than 10 Fraunhofer lines could be detected as prominent absorption lines when the wavelength resolution is higher than 1 nm/pix (30 cm−1/pix at 1000 cm−1). This result suggests that the target-free wavelength-calibration method using solar Fraunhofer lines can be applied to other spectrometers simply by observing sunlit planetary surfaces.

Does radiographic calibration affect linear radiographic measurements in a large pediatric spine registry?

Large registries are increasingly at the forefront of modern pediatric spine research, with manual, centralized, trained radiographic measurement serving as the gold standard for spine research. However, there is limited data regarding the reliability of registry measurements which may be subject to differences in radiographic calibration.We undertook this study to evaluate reliability of T1-T12 height, L1-S1 height, and coronal balance measurements for a large registry of early onset scoliosis patients. Three trained technicians from the Pediatric Spine Study Group measured 43 radiographs for T1-T12, L1-S1, and coronal balance using 3 different calibration techniques. All radiographs were AP views of patients with magnetically controlled growing rods with known diameters. The calibration techniques used a pre-export manually drawn line, a digital automatically generated computerized marker, and the diameter of a magnetically controlled growing rod.The intraclass correlation coefficient (ICC) was calculated to determine reliability. 1161 measurements were performed.For each of the three raters, coronal balance, T1-T12 height and L1-S1 height had excellent agreement regardless of the calibration technique (alpha 0.93-1.0).Among the parameters, coronal balance had the worst inter-rater reliability, whereas there was excellent interrater reliability regarding T1-T12 height and L1-S1 height (alpha 0.91-0.99). There was excellent agreement among reviewers and between the 3 different calibration techniques. While calibration using rod diameter served as the gold standard, this data shows that other standard calibration methods were adequate and achieved excellent reliability for registry radiographs.

Solubility and Taste Masked Behaviour of Cyclodextrin Molecular Inclusion Complex of Artemether

Artemether by forming a complex with β-cyclodextrine (βCD) is used to improve solubility and taste masked by the kneading method. Artemether with β-cyclodextrine complex prepared with βCD in a drug-polymer ratio 1:1 gave complete taste masking with the satisfactory result obtained in term of in-vivo and in-vitro evaluation. The standard calibration curve of Artemether shows a slope of 0.0154 and a correlation coefficient of 0.9992. Compatibility studies show that drugs are compatible with polymers. Solubility studies show that the solubility of drug are increased in Phosphate buffer 6.8 by successfully forming a complex with βCD. In-vitro dissolution results showed that a maximum of 97.05% of Artemether, drug release less than 15 minutes in formulation. The solubility of Artemether in saliva (PH 6.8) is 0.00735 mg/mL, After complex forming with β-cyclodextrin the solubility of (Artemether complex) is 0.124 mg/mL. In-vitro drug release investigation, taste masking occurs when the amount of drug substance in the dissolve medium is either not identified or is below the threshold for recognizing its flavor in the early time points (between 0 and 5 minutes). Taste masking ( Taste perception test) 11 volunteers were selected for this test, results show that complex is taste masked. Artemether with βCD complex is a hopeful method to augment the solubility, dissolution and taste masking of the drug.

Solubility and Taste Masked Behaviour of Cyclodextrin Molecular Inclusion Complex of Lumefantrin

Lumefantrine is an antimalarial drugs used especially for pediatric persons. β-Cyclodextrine is used to improve the taste of lumefantrine by forming complexes with them by the solvent evaporation method, respectively. β-Cyclodextrine prevent the release of drug in the saliva. Lumefantrine is a bitter drug. So masking of bitter taste in the formulation is a prerequisite as it improves the compliance of the patient and product value. Lumefantrine with β-cyclodextrin in a drug-polymer ratio 1:1 gave complete taste masking with a satisfactory result obtained in term of in-vivo and in-vitro evaluation visual inspection revealed that lumefantrine was yellow-crystalline. The melting point of lumefantrine was found to be 127℃, respectively. The standard calibration curve of Lumefantrine shows a slope 0.0108 and a correlation coefficient of 0.9997. Compatibility studies show that drugs are compatible with polymers. Solubility study show that the solubility of both drugs are increased in phosphate buffer 6.8 by successfully forming a complex with β-cyclodextrin. The solubility of lumefantrine in saliva (pH 6.8) is 0.408 mg/mL, after complex forming with β-cyclodextrine the solubility of (Lumefantrine complex) is 0.526 mg/mL. In an in-vitro drug release investigation, taste masking occurs when the amount of drug substance in dissolved media is either below the detection threshold for recognizing its taste or is undetectable in the early time points (between 0 and 5 minutes). Taste masking (Taste perception test) 11 volunteers are selected for this test, results show that complex is taste masked successfully. Lumefantrine with β-Cyclodextrine complex is a hopeful approach to enhance solubility, dissolution and drug taste.

Determination of Paraphenylenediamine in Hair Dyes Formulation Using Visible Spectroscopy

Paraphenylenediamine (PPD), which stands for para-phenylenediamine, is a substance commonly present in hair dyes and is a potent sensitizer and reported to have adverse effects, including skin rashes, allergies, etc. This research study introduces a newly developed and rigorously tested visible spectrophotometric technique for accurately measuring the quantity of PPD in hair dye products. The proposed method relies on a colorimetric reaction between PPD and Folin–Ciocalteu phenol (FC) reagent in the presence of sodium hydroxide (NaOH), which results in a PPD-Folin’s reagent complex that is blue in color. To create a standard calibration curve, various concentrations of PPD solutions within the range of 2 to 10 μg/mL, dissolved in 0.1N NaOH, were combined with 1.5 mL of FC reagent solution and scanned at 645 nm. The method was then used to estimate the amount of PPD in three marketed hair dye formulations. The technique demonstrated accuracy and precision, with a detection limit of 0.2445 μg/mL and a quantification limit of 0.7411 μg/mL. The level of PPD observed in three marketed formulations was between 10 to 12.5 μg/mL. During the study, it was observed that the black henna pack contained high concentrations of PPD

Absolute contents of aroma-affecting volatiles in cooked rice determined by one-step rice cooking and volatile extraction coupled with standard-addition calibration using HS-SPME/GC–MS

To quantify volatiles in cooked rice, analysis methods for one-step rice cooking and volatile extraction in a single headspace vial, combined with standard addition calibration using solid-phase microextraction and GC–MS were developed and applied to 41 rice varieties with various fragrances and palatability. The newly developed methods significantly improved the qualitative and quantitative recovery of volatiles compared with conventional methods. Among 29 aroma-affecting volatiles, the highest average contents (ng/g) were observed for nonanal (39.30), octanal (13.29), and 1-octen-3-ol (13.18); the total volatile contents of aldehyde, base, and alcohol groups were 4156, 2481, and 1739 ng/g, respectively. Fifteen rice varieties contained 2-acetyl-1-pyrroline in range of 41.37–421.70 ng/g. Although there were no linear correlations among volatiles and the Toyo taste-score, multivariate PLS-DA analysis of the volatile could discriminate between low- and high-palatability rice varieties. The results indicated the accuracy and practicality of the newly developed methods for quantifying volatiles in cooked rice.

BART: A transferable liquid chromatography retention time library for bile acids

Identification of unknown bile acids, especially the distinguishment between isomers, requires retention times of a large number of reference standards, which are often not commercially available. Meanwhile, published retention information cannot be directly transferred across labs due to the differences between liquid chromatography (LC) systems, such as different extra column volume and dwell volume. To improve this situation, a transferrable retention time library for bile acids named BART was developed. BART was composed of isocratic retention models of 272 bile acids and a software tool to predict their gradient retention times on various LC systems. The isocratic retention times of bile acids were acquired on a Waters BEH C18 column with mobile phases of acidic ammonium acetate buffer and acetonitrile, and fit to the quadratic solvent strength model (QSSM). Segmented linear gradient retention times were calculated with holdup time (t0), dwell time (tD) and actual gradient profile corrected using 21 bile acid calibration standards. In addition to the reference system where the isocratic retention times were acquired, this approach has been validated on four other LC-MS systems in four labs with two gradient methods. Average root mean square errors (RMSE) between predicted and experimental retention times were 0.052 and 0.054 min for the two gradients tested, which were 9-fold more accurate than referring to a static retention time library. The library is freely available at https://bafinder.github.io/.

A workflow for the detection of antibiotic residues, measurement of water chemistry and preservation of hospital sink drain samples for metagenomic sequencing

Hospital sinks are environmental reservoirs that harbour healthcare-associated (HCA) pathogens. Selective pressures in sink environments, such as antibiotic residues, nutrient waste and hardness ions, may promote antibiotic resistance gene (ARG) exchange between bacteria. However, cheap and accurate sampling methods to characterize these factors are lacking. To validate a workflow to detect antibiotic residues and evaluate water chemistry using dipsticks. Secondarily, to validate boric acid to preserve the taxonomic and ARG ('resistome') composition of sink trap samples for metagenomic sequencing. Antibiotic residue dipsticks were validated against serial dilutions of ampicillin, doxycycline, sulfamethoxazole and ciprofloxacin, and water chemistry dipsticks against serial dilutions of chemical calibration standards. Sink trap aspirates were used for a 'real-world' pilot evaluation of dipsticks. To assess boric acid as a preservative of microbial diversity, the impact of incubation with and without boric acid at ∼22°C on metagenomic sequencing outputs was evaluated at Day 2 and Day 5 compared with baseline (Day 0). The limits of detection for each antibiotic were: 3μg/L (ampicillin), 10μg/L (doxycycline), 20μg/L (sulfamethoxazole) and 8μg/L (ciprofloxacin). The best performing water chemistry dipstick correctly characterized 34/40 (85%) standards in a concentration-dependent manner. One trap sample tested positive for the presence of tetracyclines and sulphonamides. Taxonomic and resistome composition were largely maintained after storage with boric acid at ∼22°C for up to five days. Dipsticks can be used to detect antibiotic residues and characterize water chemistry in sink trap samples. Boric acid was an effective preservative of trap sample composition, representing a low-cost alternative to cold-chain transport.

Sustainable Coffee Empowerment in Saudi Arabia: Unleashing Potential Through Comprehensive Cupping Sessions

This paper describes a study that used coffee cupping sessions to evaluate various coffee samples from green coffee merchants in Saudi Arabia. The study focuses on attaining quality assurance, assessing suppliers, empowering people, encouraging sustainability, and differentiating the firm in the market. The study integrates theories such as Expectancy Disconfirmation Theory, Resource-Based View, Self-Determination Theory, Triple Bottom Line, and Differentiation Strategy. Qualitative and quantitative elements were blended in a mixed methods approach. Randomly chosen single origin coffee samples were procured from trustworthy vendors in Jeddah, Riyadh, and Dammam. Cupping sessions were held in accordance with the Specialty Coffee Association (SCA) standard cupping methodology, and experienced participants rated the samples. The results and analysis included an overview of the cupping sessions, sensory evaluation data analysis, a comparison of samples from various suppliers, and the identification of important taste profiles and attributes. Employee empowerment was also stressed in the study through coffee evaluation, which included training and educational activities, the impact on participants' knowledge and abilities, and their feedback and reflections. Furthermore, the study looked at the consequences of sustainability, focusing on supporting sustainable practices, strengthening partnerships with suppliers, and improving quality to increase market competitiveness. The cupping session technique was described in depth in the professional report, which included randomization and sample division, roasting and resting methods, standardized roasting procedures, calibration and perception sessions, blind cupping, transparency measures, and post-cupping session communication

An Integrated ddPCR Lab-on-a-Disc Device for Rapid Screening of Infectious Diseases.

Digital droplet PCR (ddPCR) is a powerful amplification technique for absolute quantification of viral nucleic acids. Although commercial ddPCR devices are effective in the lab bench tests, they cannot meet current urgent requirements for on-site and rapid screening for patients. Here, we have developed a portable and fully integrated lab-on-a-disc (LOAD) device for quantitively screening infectious disease agents. Our designed LOAD device has integrated (i) microfluidics chips, (ii) a transparent circulating oil-based heat exchanger, and (iii) an on-disc transmitted-light fluorescent imaging system into one compact and portable box. Thus, droplet generation, PCR thermocycling, and analysis can be achieved in a single LOAD device. This feature is a significant attribute for the current clinical application of disease screening. For this custom-built ddPCR setup, we have first demonstrated the loading and ddPCR amplification ability by using influenza A virus-specific DNA fragments with different concentrations (diluted from the original concentration to 107 times), followed by analyzing the droplets with an external fluorescence microscope as a standard calibration test. The measured DNA concentration is linearly related to the gradient-dilution factor, which validated the precise quantification for the samples. In addition to the calibration tests using DNA fragments, we also employed this ddPCR-LOAD device for clinical samples with different viruses. Infectious samples containing five different viruses, including influenza A virus (IAV), respiratory syncytial virus (RSV), varicella zoster virus (VZV), Zika virus (ZIKV), and adenovirus (ADV), were injected into the device, followed by analyzing the droplets with an external fluorescence microscope with the lowest detected concentration of 20.24 copies/µL. Finally, we demonstrated the proof-of-concept detection of clinical samples of IAV using the on-disc fluorescence imaging system in our fully integrated device, which proves the capability of this device in clinical sample detection. We anticipate that this integrated ddPCR-LOAD device will become a flexible tool for on-site disease detection.

Direct quantification of 1,1-dimethylhydrazine transformation products in sandy soil by thermal desorption gas chromatography – Tandem mass spectrometry

The use of rocket propellants based on unsymmetrical dimethylhydrazine (UDMH) in launch vehicles and boosters poses a threat to the environment due to the release of UDMH into soils and its subsequent transformation into a variety of nitrogen-containing compounds, including toxic and carcinogenic ones. The present proof-of-concept study proposes a novel approach to screening and simultaneous determination of a wide range of nitrogen-containing UDMH transformation products in soils, based on direct thermal desorption (TD) as a solvent-free sample extraction technique coupled with GC–MS/MS analysis.Optimal TD conditions consisting of the incubation of 5 mg sample at 300 °C for 30 min, allowed attaining high recoveries (80–100 %) for the most tested compounds and developing the method for quantification of fifteen acyclic and heterocyclic UDMH transformation products in sandy soil with LODs of 0.3–7.8 µg kg−1. The matrix matched calibration and introduction of the internal standard (pyridine‑d5) ensured high accuracy (75–100 %) and precision (RSD < 15 %) of the analysis. Along with high sensitivity and accuracy, the main advantages of the developed method are exceptional simplicity of sample preparation, low amount of toxic sample, high automation degree, and environmental friendliness.The developed method was successfully tested in the analysis of seven real soil samples taken from the Baikonur cosmodrome operational zone (Kazakhstan). The use of TD-GC–MS analysis in full spectrum scanning mode allowed for tentative identification of 62 UDMH transformation products including several compounds not yet described in the literature.

Method Development for Quantitative Analysis of Polycyclic Aromatic Hydrocarbons, Nitrogen Heterocycles and Sulfur Heterocycles in Crude Oils Using Quadrupole Time-of-Flight Mass Spectrometry

An oil spill is a catastrophic event that results in various toxic polycyclic aromatic hydrocarbons (PAHs) entering the environment. Polycyclic aromatic nitrogen heterocycles (PANHs) are more toxic to the environment than their parent PAHs. The high cost and paucity of available PANH standards, the lower abundance of PANHs relative to PAHs, and the difficult separation due to co-elution with PAHs have all contributed to the scarcity of related published literature on the determination of these compounds. To overcome these challenges, a new quantitative method has been successfully developed and validated for the inclusion of 113 polycyclic aromatic carbon (PAC) compounds in a single injection. The 113 compounds consist of PAHs, nitrogen heterocycles, sulfur heterocycles, and alkylated equivalents. Distinct separation of the PANHs and their alkylated counterparts (APANHs) from PAHs was achieved using a gas chromatography quadrupole time-of-flight (GC-QToF) mass spectrometer. The instrument resolved compounds by the high-resolution extraction of monoisotopic masses, allowing response correction factors (RCFs) to be determined from available PANH standards and to calculate concentrations from PAH calibration standards. The developed method was applicable to crude oil samples, generating concentrations of PANHs and relevant information on compound stability for use in oil spill forensics investigation. Development of this practical PAC method provides a powerful tool for screening toxic contaminants, assessing environmental impact, and monitoring recovery following an oil spill.

A Non-Matrix-Matched Calibration Method for In Situ Major and Trace Element Analysis of Scheelite by Nanosecond LA-ICP-MS.

In this work, a reliable and robust in situ non-matrix-matched calibration method is proposed for element composition determination in scheelite samples. With external calibration against the silicate glass standard reference material NIST SRM 610, the concentrations of both major elements (Ca and W) and trace elements (Si, Fe, Mo, Y, rare earth elements, etc.) in scheelite are determined using an ArF 193 nm excimer nanosecond laser ablation-inductively coupled plasma mass spectrometer (LA-ICP-MS). Here, the ablation was performed by hole drilling under a helium (He) environment using a laser spot size of 35 μm and a laser repetition of 5 Hz, and the aerosols were then transported to a quadrupole ICP-MS by a mixture of He and make-up gas argon (Ar) with a total gas flow rate of 1.6 L/min. Results showed that there was no apparent matrix effect between the NIST SRM 610 and scheelite by this proposed method. With internal standardization against W, the obtained concentrations of CaO and WO3 were found to yield an average matrix CaO/WO3 mass fraction ratio of 0.245 (2σ = 0.003, n = 19), which agreed well with the value of 0.243 (2σ = 0.002, n = 15) from electron probe microanalysis (EPMA). Furthermore, the accuracy of trace element analyses with this proposed non-matrix-matched calibration in situ method was evaluated by comparing the concentration results with those from bulk analysis by solution nebulizer ICP-MS (SN-ICP-MS). It was found that the quantification results from LA-ICP-MS and SN-ICP-MS were comparable, in particular showing a relative concentration bias of the total ∑REE+Y contents of less than 2%. This confirmed that scheelites can be accurately analyzed in situ by LA-ICP-MS without matrix-matched calibration standards. By using this developed in situ method, the element compositions in a series of scheelite samples from different W-associated deposits in China were successfully quantified, promising further genetic process investigation and associated geologic activities of the polymetallic resources.

Determination of psilocybin and psilocin content in multiple Psilocybe cubensis mushroom strains using liquid chromatography – tandem mass spectrometry

A method for clinical potency determination of psilocybin and psilocin in hallucinogenic mushroom species Psilocybe cubensis was developed using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Five strains of dried, intact mushrooms were obtained and analyzed: Blue Meanie, Creeper, B-Plus, Texas Yellow, and Thai Cubensis. An extraction protocol was developed; this included an evaluation of sample milling technique, extraction solvents, and recovery/stability. Reversed phase chromatography on fused-core particle phases was developed for the determination of the two analytes using internal standard calibration with deuterated isotopologues of each analyte. The separation takes less than 5 min. Matrix effects were investigated by comparing signal response of calibration samples in neat solution and several mushroom matrices; no significant matrix effects were observed. The limit of detection for psilocybin was 1.5 ng/mL (1.5 pg on-column; 300 ng/g mushroom) and for psilocin was 0.15 ng/mL (0.15 pg on-column; 30 ng/g mushroom) using a Shimadzu LCMS-8050 triple quadrupole mass spectrometer. Assessment of the accuracy and precision of the method indicated percent error and RSD were <6 % at all concentration levels. Three whole, intact mushrooms from each strain were analyzed individually to obtain average content differences both between strains and between mushrooms of the same strain. From most to least potent, the study found that the average total psilocybin and psilocin concentrations for the Creeper, Blue Meanie, B+, Texas Yellow, and Thai Cubensis strains were 1.36, 1.221, 1.134, 1.103, and 0.879 % (w/w), respectively. A subset of these mushrooms was also tested in a separate non-affiliated laboratory, and the results were comparable between the two laboratories. Results from the secondary laboratory showed improved precision when multiple mushrooms were homogenized together, prior to extraction.

"Portrait" of a patient with vitamin D deficiency in the blood (according to one of the clinics in Tyumen)

Introduction. This work is devoted to the analysis of parameters (social and laboratory data) that allowed creating a profile of a patient with vitamin D deficiency. It is known that vitamin D is not only involved in the regulation of calcium and phosphorus metabolism, but also has extra-bone effects. However, disorders in these processes do not always have specific manifestations that make the doctor and the patient think about vitamin D deficiency. The study found that 64 % of the patients had vitamin D deficiency in their blood. Considering the above, a doctor of any specialty should remember about the ambiguous clinical picture of vitamin D deficiency and have a certain alertness when diag nosing this condition. Objective: to evaluate the social and laboratory data available in the medical record compiling a profile of a patient with vitamin D deficiency. Materials and methods. The work was carried out on the basis of a multidisciplinary clinic of the Tyu men State Medical University in the period of 2019–2020. The study included 231 medical records. The D3 content in the blood was determined by laboratory immunological method of quantitative determination of the D3 level in vitro. The ELISA method was used. The results were calculated based on a standard calibration curve. Results. In the course of the study, it was possible to make a profile of a patient with vitamin D defi ciency in the blood. Conclusion. When managing a patient with a particular pathology, determining the level of vitamin D in the blood plays an essential role as this, among other things, will allow having an individual approach to patient treatment and examination.

Создание калибровочных стандартов для мониторинга минимальной остаточной болезни методом количественной ПЦР с обратной транскрипцией при ОМЛ с t(10;11)(p12.31;q14.2)/ PICALM::MLLT10

Хромосомная транслокация t(10;11)(p12.31;q14.2) с образованием химерного гена PICALM::MLLT10 является редкой рекуррентной перестройкой при некоторых типах острых лейкозов, включая острый миелоидный лейкоз (ОМЛ). Она ассоциирована с неблагоприятным прогнозом и в настоящее время включена в риск-стратификацию ОМЛ у детей, получающих терапию по протоколу ОМЛ-MRD-2018, в качестве маркера высокого риска. Экспрессия PICALM::MLLT10 может использоваться в качестве мишени для мониторинга минимальной остаточной болезни (МОБ) методом количественной ПЦР с обратной транскрипцией. Однако в силу редкости и гетерогенности данной перестройки для нее отсутствуют коммерчески доступные калибровочные стандарты. Целью данной работы явилось создание калибровочных стандартов для оценки экспрессии химерного гена PICALM::MLLT10 методом клонирования в плазмидный вектор. Были заклонированы 4 фрагмента PICALM::MLLT10 с различными локализациями точек разрыва в гене MLLT10 (интроны 4, 6, 9 и 10) и локализацией точки разрыва в гене PICALM в интроне 19. Для каждого калибровочного стандарта было показано наличие корректной вставки методом секвенирования по Сэнгеру и отсутствие мутаций в сайтах посадки праймеров и проб для количественной ПЦР. Чувствительность примененных в настоящей работе систем детекции составила 10 копий в реакции в ПЦР с плазмидными стандартами и 1/10 000 в 10-кратных лимитирующих разведениях. Значение углового коэффициента для калибровочных стандартов составило 3,4±0,05, свободного коэффициента – 34,7±1,24, значение R2 превышало 0,95 во всех случаях, коэффициент вариации для образцов с минимальным содержанием молекул-мишеней составил 1,12±0,43%, что соответствует международным рекомендациям для систем детекции и количественных стандартов, применяемых в мониторинге МОБ методом ОТ-ПЦР в режиме реального времени. The chromosomal translocation t(10;11)(p12.31;q14.2) leading to PICALM::MLLT10 fusion gene formation is a rare recurrent rearrangement in some types of acute leukemia, including acute myeloid leukemia (AML). It is associated with a poor prognosis and is currently included in the risk stratification of AML in children receiving therapy according to the AML-MRD-2018 protocol as a high-risk marker. PICALM::MLLT10 expression can be used as a target for minimal residual disease (MRD) monitoring by quantitative reverse transcription-PCR. However, due to the rarity and heterogeneity of this rearrangement, there are no commercially available calibration standards for it. The goal of this work was to create calibration standards for assessing the expression of the chimeric gene PICALM::MLLT10 by cloning into a plasmid vector. Four fragments of PICALM::MLLT10 were cloned with different localizations of breakpoints in the MLLT10 gene (introns 4, 6, 9 and 10) and localization of the breakpoint in the PICALM gene in intron 19. For each calibration standard, the presence of a correct insert was demonstrated by Sanger sequencing and the absence of mutations in the primer binding sites and probes for quantitative PCR. The sensitivity of the detection systems used in this work was 10 copies in a PCR reaction with plasmid standards and 1/10000 in 10-fold limiting dilutions. The slope coefficient for calibration standards was 3.4±0.05, y-intercept was 34.7±1.24, the R2 value exceeded 0.95 in all cases, the coefficient of variation for samples with a minimum content of target molecules was 1.12±0.43%, which corresponds to international recommendations for systems detection and quantitative standards used in real-time RT-PCR monitoring of MRD.

Quantifying uncertainty in analysis of shockless dynamic compression experiments on platinum. I. Inverse Lagrangian analysis

Absolute measurements of solid-material compressibility by magnetically driven shockless dynamic compression experiments to multi-megabar pressures have the potential to greatly improve the accuracy and precision of pressure calibration standards for use in diamond anvil cell experiments. To this end, we apply characteristics-based inverse Lagrangian analysis (ILA) to 11 sets of ramp-compression data on pure platinum (Pt) metal and then reduce the resulting weighted-mean stress–strain curve to the principal isentrope and room-temperature isotherm using simple models for yield stress and Grüneisen parameter. We introduce several improvements to methods for ILA and quasi-isentrope reduction, the latter including calculation of corrections in wave speed instead of stress and pressure to render results largely independent of initial yield stress while enforcing thermodynamic consistency near zero pressure. More importantly, we quantify in detail the propagation of experimental uncertainty through ILA and model uncertainty through quasi-isentrope reduction, considering all potential sources of error except the electrode and window material models used in ILA. Compared to previous approaches, we find larger uncertainty in longitudinal stress. Monte Carlo analysis demonstrates that uncertainty in the yield-stress model constitutes by far the largest contribution to uncertainty in quasi-isentrope reduction corrections. We present a new room-temperature isotherm for Pt up to 444 GPa, with 1-sigma uncertainty at that pressure of just under ±1.2%; the latter is about a factor of three smaller than uncertainty previously reported for multi-megabar ramp-compression experiments on Pt. The result is well represented by a Vinet-form compression curve with (isothermal) bulk modulus K0=270.3±3.8 GPa, pressure derivative K0′=5.66±0.10, and correlation coefficient RK0,K0′=−0.843.

Simultaneous extraction and quantitative analysis of S-Methyl-l-Cysteine Sulfoxide, sulforaphane and glucosinolates in cruciferous vegetables by liquid chromatography mass spectrometry

Sulfur containing compounds including glucosinolates (GLS), sulforaphane (SFN) and S-methyl-l-cysteine sulfoxide (SMCSO) have been proposed to be partly responsible for the beneficial health effects of cruciferous vegetables. As such, greater understanding of their measurements within foods is important to estimate intake in humans and to inform dietary intervention studies. Herein is described a simple and sensitive method for simultaneous analysis of 20 GLS, SFN and SMCSO by liquid chromatography mass spectrometry. Analytes were effectively retained and resolved on an Xbridge C18 column. Detection can be achieved using high resolution or unit resolution mass spectrometry; the latter making the method more applicable to large studies. Quantitative analysis using calibration standards was demonstrated for 10 GLS, SFN and SMCSO. A further 10 GLS were tentatively identified using high resolution mass spectrometry. The use of surrogate GLS standards was shown to be unreliable, with closely related GLS displaying significantly different ionisation efficiencies.

HPLC-LED-Induced Fluorescence Analysis of Tear Fluids: An Objective Method for Primary Open Angle Glaucoma Diagnosis

PurposeThe study showcased the application of the lab-assembled HPLC-LED-IF system to analyze proteins in tear fluid samples collected from individuals diagnosed with primary open-angle glaucoma (POAG).MethodsClinical application of the said technique was evaluated by recording chromatograms of tear fluid samples from control and POAG subjects and by analyzing the protein profile using multivariate analysis. The data analysis methods involved are principal component analysis (PCA), Match/No-Match, and artificial neural network (ANN) based binary classification for disease diagnosis.ResultsMahalanobis distance and spectral residual values calculated using a standard calibration set of clinically confirmed POAG samples for the Match/No-Match test gave 86.9% sensitivity and 81.8% specificity. ANN with leaving one out procedure has given 87.1% sensitivity and 81.8% specificity.ConclusionThe results of the study revealed that the utilization of a 278 nm LED excitation in the HPLC system offers good sensitivity for detecting proteins at low concentrations allowing to obtain reliable protein profiles for the diagnosis of POAG.

Dilute-and-shoot ICPMS quantification of V, Ni, Co, Cu, Zn, As, Se, Ag, Cd, Ba, and Pb in fruit juices based on matrix overcompensation calibration.

Analysis of V, Co, Ni, Cu, Zn, As, Se, Ag, Cd, Ba, and Pb in fruit juices was performed by inductively coupled plasma mass spectrometry (ICPMS) after simple 50-fold dilution in 1% (v/v) HNO3-0.5% (v/v) HCl-5% (v/v) ethanol. Ethanol was added to overwhelm native organic components and dominate matrix effects. A universal calibration curve was built based on a likewise treated reagent standard series. This new matrix overcompensation calibration (MOC) strategy was developed to effectively compensated formatrix effects of carbon origin and achieved quantitative (92.5-118.8%) recoveries comparable to those by standard addition calibration (92.1-117.8%) and microwave-aided digestion (99.3-116.8%). The LODs were 0.528, 0.204, 0.195, and 2.07ngmL-1 for toxic elements As, Cd, Pb, and Ni, respectively, adequate for their regulatory monitoring. Ge, Rh, Tb, and Ir were used as internal standards. MOC renders a calibration curve universally applicable to any clear fruit juices of diversified crop, geographic, and manufacturer origins resulting in cost saving and enhanced productivity.