Sample Preparation Method Research Articles

Top dusted adhesive tape sample preparation method for the X-ray diffraction analysis of small powder sample volumes with the Bragg–Brentano setup

When performing X-ray diffraction (XRD) analysis of small powder sample volumes with the Bragg–Brentano setup, choosing an appropriate sample preparation method is important. This work provides practical information on the conventional top dusted sample preparation method and the top dusted adhesive tape sample preparation method. In the latter, the powder of interest is dusted onto an adhesive tape. Because adhesive tapes contribute to the measured XRD signal as background noise, care must be taken when using a particular adhesive tape. This work assists XRD users in selecting the most suitable adhesive tape for their purpose by providing the diffractograms (Cu Kα1 radiation) of 13 commercially available adhesive tapes used for office and packing applications. In general, tapes with a matte acetate or polyvinyl chloride backing are recommended as they cause a comparatively low background signal without high-intensity reflections.

A novel 3D-printed sample preparation method for benzodiazepine quantification in human serum.

Benzodiazepine abuse remains a significant public health concern. Current sample preparation methods for benzodiazepine analysis from human serum often involve complex procedures that require large sample volumes and extensive organic solvent use. To address these limitations, this study presents a novel and efficient sample preparation method utilizing 3D-printed sorbent devices. The 3D-printed devices, fabricated from a thermoplastic composite incorporating C18-modified silica, demonstrated exceptional performance in extracting benzodiazepines from human serum. The method was optimized and validated according to ICH guidelines, ensuring its reliability for quantitative benzodiazepine analysis. Notably, the method required minimal sample and solvent volumes, eliminating the need for protein precipitation, evaporation, and reconstitution. This novel sample preparation approach offers significant advantages over traditional methods, providing a more efficient and environmentally friendly solution for benzodiazepine analysis. The versatility of 3D printing allows for the customization of sorbent devices for various analytes and matrices, expanding the potential applications of this method. Coupled with a rapid and robust LC-MS method optimized with DryLab, this approach presents a valuable and sensitive tool for benzodiazepine monitoring in clinical and toxicological settings.

Upgradation in biosensor: An emerging paradigm for advanced biomedical application

With the advent of the development of biosensor devices, scientists are more focused on emerging new biosensor technologies and tools. The existing application of these devices in fields of biomedicine, drug discovery, food safety, and environmental monitoring get remarkable importance. Till today diagnostic tests especially for early detection and monitoring of cancer and anticancer chemotherapeutic agents bearing major challenges despite their simplicity. This review aims to highlight the use of biosensors as diagnostic means along with their future applicability. In the past various achievement in the development of biosensors opened a new gateway for researchers to further modify biosensing elements to enhance their detection ability particularly in cancer that requires continued development and validation of biomarkers and development of ligands for those biomarkers, as well as continued development of sample preparation methods and multichannel biosensors able to analyze many cancer markers simultaneously. The future for biosensor is promising and bright however require a determined, cost-effective and multi-disciplinary approach to bring out biosensor system from research and development laboratory to market place.

Influence of Sample Preparation Method on Total Dietary Fiber Content Measurement of Low Carbohydrate Bread Containing High Levels of Cross‐Linked Phosphorylated RS4 Wheat Starch

ABSTRACTBackground and ObjectivesCross‐linked phosphorylated (CLP) RS4 wheat starch has been approved as a dietary fiber source by FDA because it can reduce insulin levels following a meal containing a carbohydrate that raises blood glucose levels. It has been successfully formulated in many high‐fiber and low‐carbohydrate bread products. This study aimed to determine the influence of sample preparation methods on the total dietary fiber content of these fiber‐fortified bread products during the assay with AOAC Method 991.43. In addition, the effects of CLP RS4 wheat starch on the morphology, crystallinity, and thermal properties of the bread were investigated.FindingsThe total dietary fiber content of breads formulated with unmodified wheat starch or CLP RS4 wheat starch was influenced by the sample preparation methods (drying, grinding, and sieving). Preferred methods are forced air oven over freeze‐drier and vacuum oven; coffee grinder over mortar and pestle; and 35‐mesh over 70‐mesh sieve. The proposed method excludes influence of operator and shows more repeatable results. CLP RS4 wheat starch in the bread was gelatinized and had restricted swelling, reduced enthalpy, and reduced crystallinity.ConclusionsThe proposed sample preparation method generated from this study involves drying the bread in a forced air oven at 40°C, particle size reduction using a coffee grinder, and screening the ground samples in a 35‐mesh sieve before total dietary fiber assay using AOAC Method 991.43.Significance and NoveltyOur study provided new and useful information on sample preparation methodology for total dietary fiber measurement of high‐fiber or low‐carbohydrate bread products containing CLP RS4 wheat starch. This methodology has potential applications in many high‐fiber flour‐based products other than bread.

The practice of using varios methods of sample preparation for the X-ray fluorescence determination of macroelements in geological rocks, soils and sediments

To determine the content of macronutrients in rocks, soils and sediments, three methods of sample preparation were used by the X-ray fluorescence method: pressing powder samples; fusing in gold-platinum crucibles to obtain glass discs; fusing on graphite substrates followed by melting and tablet pressing. A detailed description of each of the methods used is provided. For fusion in gold-platinum crucibles, a ready-made LITHIUM BORATE FLUX (from the company CHEMPHYS) was used, its composition: Li2B4O7 (67%) + LiBO2 (33%) + alloying additive LiBr. For fusion on graphite substrates, powders of lithium tetraborate and metaborate in a ratio of 2:1, dried at 600°C. Random errors of the analysis are estimated: errors of sample preparation and intensity measurement. It is shown that in all cases, the total errors do not exceed the permissible level when conducting mass analyses of the III category. Standard samples of rocks, sediments and soils were used to obtain the grades, The metrological characteristics of X-ray fluorescence analysis techniques using each of the three sample preparation methods are presented. The residual standard deviations of the calibration dependencies are compared. The correctness of the analysis results was verified on three standard samples. The advantages and disadvantages of each sample preparation method are noted. The use of the simplest, cheapest and express method of pressing tablets from the source material is possible only if the content of elements in the analyzed samples does not exceed the limits of the calibration dependencies of this technique. The method of fusion in gold-platinum crucibles requires preliminary information about the chemical composition of the analyzed sample. The method of fusing on graphite substrates (followed by melting and pressing the tablet) is more versatile and has no restrictions on the composition of the fused sample. It is shown that the systematic errors of the method for pressed tablets exceed the errors of the methods using the second and third techniques. The work was performed on the AXIOS Advanced wave dispersive X-ray fluorescence spectrometer (PANalytical B.V., Holland).

A Fast, High-Sensitivity 96-Well Plate-Based MICROFASP Method for Processing Low Microgram Proteomics Sample within 1.5 h.

A rapid, sensitive, and high-throughput sample preparation method is of paramount significance for proteomics analysis. Here, we report a fast, high-sensitivity MICROFASP method that is capable of completing sample preparation within 1.5 h, enhancing the throughput by over 13 times compared to the previous reports. Protein digestion time was significantly cut from 17 h to 20 min in a limited volume. Simultaneous reduction and alkylation occurred within 30 min. The label-free quantitation intensities of proteins from the fast and conventional MICROFASP methods were highly correlated (r = 0.91), validating the reliability of the fast-MICROFASP method. When starting with 1 μg of K562 cell lysate, the fast-MICROFASP method identified over 6 times more protein groups and 19 times more peptides than did the iST method. A 96-well plate-based version was developed to process 8 brain tissue samples from APP/PS1 transgenic mice in parallel. Averagely, with only 1 μg of protein lysate, 2826 protein groups (n = 8, RSD = 0.7%) and 12,972 peptides (n = 8, RSD = 1.5%) were identified from each sample. Amyloid-beta protein was successfully identified as a highly expressed protein, which shows its potential for detecting diagnostic markers and proteome profiling with low-microgram samples. We anticipate the high-sensitivity 96-well plate-based fast-MICROFASP method will have wide application in high-throughput and rapid preparation of large cohorts of low-microgram samples (e.g., clinical biopsy) for comprehensive proteome profiling. Data are available via ProteomeXchange with the identifier PXD053720.

Gel-Based Sample Fractionation with SP3-Purification for Top-Down Proteomics.

Precise prefractionation of proteome samples is a potent method for realizing in-depth analysis in top-down proteomics. PEPPI-MS (Passively Eluting Proteins from Polyacrylamide gels as Intact species for MS), a gel-based sample fractionation method, enables high-resolution proteome fractionation based on molecular weight by highly efficient extraction of proteins from polyacrylamide gels after SDS-PAGE separation. Thereafter it is essential to effectively remove contaminants such as CBB and SDS from the PEPPI fraction prior to mass spectrometry. In this study, we developed a complete, robust, and simple sample preparation workflow named PEPPI-SP3 for top-down proteomics by combining PEPPI-MS with the magnetic bead-based protein purification approach used in SP3 (single-pot, solid-phase-enhanced sample preparation), now one of the standard sample preparation methods in bottom-up proteomics. In PEPPI-SP3, proteins extracted from the gel are collected on the surface of SP3 beads, washed with organic solvents, and recovered intact with 100 mM ammonium bicarbonate containing 0.05% (w/v) SDS. The recovered proteins are subjected to mass spectrometry after additional purification using an anion-exchange StageTip. Performance validation using human cell lysates showed a significant improvement in low-molecular-weight protein recovery with a lower coefficient of variation compared to conventional PEPPI workflows using organic solvent precipitation or ultrafiltration.

Evaluating sulfur-impurity removal and modern carbon contamination in different preparation methods for radiocarbon dating of soil samples by accelerator mass spectrometry

Abstract Radiocarbon (14C) dating of soil samples by accelerator mass spectrometry (AMS) has been proven useful for studying carbon (C) cycling in terrestrial ecosystems. However, this application has two primary difficulties in sample preparation: inhibition of graphite formation due to sulfur (S)-containing impurities and contamination of samples with modern C (MC). Herein, we evaluated these effects using three sample preparation methods (silver foil, silver wire, and Sulfix) by conducting AMS-14C measurements of a 14C-dead charred wood and S-rich soil samples. The preparation methods were all successful in graphite formation and AMS-14C measurement for soil samples with an organic S content <6.9 wt%. The methods showed different percent modern carbon (pMC) values from 0.16% to 0.64% for the 14C-dead sample. The results also revealed that across different methods, MC contamination can be significantly reduced by applying two-step procedure (combustion and subsequent reaction to remove S-containing impurities) during sample preparation. The three methods had a negligible influence on determining the 14C age for samples that were at least younger than 12,000 yr BP. As the 14C ages of the soil samples are typically younger than 12,000 yr BP, any method explored in this study can be employed for 14C dating with sufficient accuracy for application to C cycle studies.

Cryo-electron tomography pipeline for plasma membranes

Cryo-electron tomography (cryoET) provides sub-nanometer protein structure within the dense cellular environment. Existing sample preparation methods are insufficient at accessing the plasma membrane and its associated proteins. Here, we present a correlative cryo-electron tomography pipeline optimally suited to image large ultra-thin areas of isolated basal and apical plasma membranes. The pipeline allows for angstrom-scale structure determination with subtomogram averaging and employs a genetically encodable rapid chemically-induced electron microscopy visible tag for marking specific proteins within the complex cellular environment. The pipeline provides efficient, distributable, low-cost sample preparation and enables targeted structural studies of identified proteins at the plasma membrane of mammalian cells.

Energized dispersive guided extraction (EDGE): a promising tool for analytical chemistry applications.

Analytical chemistry demands precise sample preparation methods to ensure accurate qualitative and quantitative determinations, especially those capable of clean-up and preconcentration of target analytes. Extraction plays a crucial role in enhancing the selectivity and sensitivity of analytical procedures. Thus, Energized Dispersive Guided Extraction (EDGE) has emerged as an innovative alternative to traditional methods, such as Soxhlet, maceration, and percolation, as well as modern techniques like Accelerated Solvent Extraction (ASE), Supercritical Fluid Extraction (SFE), and Microwave or Ultrasound Assisted Extraction (MAE and UAE). By combining dispersive solid-phase extraction (DSPE) with pressurized liquid extraction (PLE), this technique offers distinct advantages, including faster extraction times, reduced solvent consumption, and improved reproducibility. Its automated system also minimizes operator intervention, ensuring greater precision and optimizing workflows. However, there is a significant gap in studies exploring its various potential applications and perspectives. In this scenario, this is the first paper to broaden the range of applications of the EDGE system by exploring its use in food, pharmaceutical, and environmental areas. These sectors benefit from EDGE's ability to preserve analyte integrity, increase extraction yields, and promote environmentally sustainable practices. Moreover, EDGE's flexibility and efficiency make it a powerful tool for addressing emerging challenges, such as detecting contaminants in complex matrices and complying with stricter regulatory standards. Expanding EDGE's applications in these areas underscores its potential to transform analytical processes, driving advancements in food safety, pharmaceutical quality control, and environmental monitoring.

Identification of Legionella by MALDI Biotyper through three preparation methods and an in-house library comparing phylogenetic and hierarchical cluster results

The identification and typing of bacteria are very expensive and time-consuming due to their growth times, and the expertise needed. MALDI-TOF MS represents a fast technique, reproducible with molecular approaches. This technique is still poorly applied in Legionella surveillance with estimation occurring only at the genus level. The aim of this study was to compare three sample preparation methods: direct smear (DS), extended direct smear (EDS), and full extraction (E), using MALDI Biotyper, developing an in-house library. Moreover, Hierarchical cluster analysis (HCA) was compared to mip and rpoB gene sequencing. The dataset was composed of 104 isolates belonging to six Legionella species. The isolates were identified with a sensitivity of 97.11% for DS, 98.08% for EDS, and 95.19% for E. The error rates were 2.88% for DS, 1.92% for EDS, and 4.90% for E, with no significant differences among them. The HCA confirmed the relationship among the isolates reported in the phylogenetic trees. An improvement in sensitivity was obtained using an in-house library. The results suggest the use of a fast and inexpensive DS method, combined with instrument and in-house library for routine Legionella surveillance. HCA analysis could be useful for screening isolates, before undertaking expensive and time-laborious molecular techniques.

Biomagnetic monitoring of urban atmospheric pollution: A review of magnetic signatures from different types of plants.

Biomagnetic monitoring has rapidly emerged as a valuable tool in urban atmospheric pollution (UAP) assessment due to its high spatial resolution, complementing traditional monitoring systems. This review systematically elucidates the principles of plant dust retention and the factors influencing it, while also reviewing the advancements in global research on UAP monitoring through the magnetic properties of various plant species. We provide a comprehensive analysis of the current applications of biomagnetic monitoring in UAP and identify critical challenges, including species-specific monitoring discrepancies, complex pollution sources, and non-standardized sample preparation methods. Additionally, we propose future research directions to address these challenges and enhance the efficacy of biomagnetic monitoring in UAP.

Amoxicillin Concentrations in Samples of Musculoskeletal Infections

Amoxicillin is a frequently used beta‐lactam antibiotic and at the same time a fragile analyte. Aim of this work was to investigate the sensitive properties of amoxicillin in biological samples and validate a quantification method for amoxicillin in soft tissue samples. Simple protein precipitation and 2D‐High performance liquid chromatography coupled to tandem mass spectrometry was used for sample preparation and analysis. The formation of a covalent amoxicillin methanol‐adduct could be observed in varying extents depending on matrix and sample preparation method. In‐Source Products (ISPs) were investigated in plasma, soft tissue, and water and no differences could be observed. We successfully validated a method for amoxicillin determination in tissue samples using the sum of 2 ISPs of amoxicillin for quantification. 37 study samples of different deep‐seated infections could be analyzed. Measured amoxicillin concentrations ranged from below 0.5 mg/kg up to 87 mg/kg. Concentrations in abscesses were lower than in other infections.

Optimization and Experimental Study of Penetration Depth of Pocket Penetrometer

The pocket penetrometer has become an ideal tool for the rapid testing of soil parameters in the field due to its portability, ease of use, and cost effectiveness, but there is still room for improvement in its accuracy. In this study, a penetration test on red clay was investigated by increasing the penetration depth and combining the self-designed penetrometer, sample preparation method, and data analysis method. The improved sample preparation technique performs well in improving sample uniformity and preparation efficiency. The quantitative processing of the penetration curves improves the objectivity and accuracy of the test results. The penetration test results of various reshaped soils showed that the relationships between liquidity index, shear strength, and penetration resistance were less affected by the liquid–plastic interface. In addition, the penetration test results of remolded soils and in situ soils showed that the empirical formulas are still informative in evaluating the shear strength of soils despite the differences in the test soils.

Migration of toxic elements from recycled paper food contact materials to food simulants: compatibility and influence of sample preparation methods

Migration of toxic elements from recycled paper food contact materials to food simulants: compatibility and influence of sample preparation methods

Exploring the Uniformity of MICP Solidified Fine Particle Silt with Different Sample Preparation Methods

Exploring the Uniformity of MICP Solidified Fine Particle Silt with Different Sample Preparation Methods

Equisetum alkaloids degradation in biogas fermentation of E. palustre contaminated plant material.

The current expansion of Equisetum palustre in wetlands across the Northern Hemisphere has led to an increase in reports of adverse effects in livestock. In light of the limited reduction potential of toxic Equisetum alkaloids through feed conservation measures, it is essential to identify effective strategies to manage E. palustre infested biomass. This study examined the impact and efficacy of biomethanization processes on the biodegradation of Equisetum alkaloids. To monitor the biodegradation in such complex matrices, a sample preparation method and a high-performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS/MS) method were established. The developed method was successfully employed to quantify the total residual content of Equisetum alkaloids in fermentation residues following biomethanization in batch experiments. It was demonstrated that under such conditions a degradation greater than 95 % was achieved. The biomethanization process described here is currently the most effective biological treatment for reducing Equisetum alkaloids. It is notable that the methane yields remained largely unaffected, indicating that field cuttings contaminated with E. palustre can be safely and economically processed via biomethanization which would be a key prerequisite for a circular bioeconomy.

An Open-source Python Tool for Traction Force Microscopy on Micropatterned Substrates.

Cell-generated forces play a critical role in driving and regulating complex biological processes, such as cell migration and division and cell and tissue morphogenesis in development and disease. Traction force microscopy (TFM) is an established technique developed in the field of mechanobiology used to quantify cellular forces exerted on soft substrates and internal mechanical tissue stresses. TFM measures cell-generated traction forces in 2D or 3D environments with varying mechanical and biochemical properties. This technique involves embedding fiducial markers in the substrate, imaging substrate deformations caused by the cells, and using mathematical models to infer forces. This protocol compiles procedures from various previously published studies and software packages and describes how to perform TFM on 2D micropatterned substrates. Although not the focus of this protocol, the methods and software packages shown here also allow to perform monolayer stress microscopy (MSM), a method to calculate internal mechanical stress within the cells by modeling them as a thin plate with linear and homogeneous material properties. TFM and MSM are non-invasive methods capable of yielding spatially and temporally resolved force and stress maps with high throughput. As such, they enable the generation of rich datasets, which can provide valuable insights into the roles of cell-generated forces in various physiological and pathological processes. Key features • TFM and MSM protocol for 2D micropatterned polyacrylamide substrates, from sample preparation over imaging to data analysis with provided code. • Sample preparation method is based on Tseng et al. [1]. • TFM analysis is done with Python custom code and is optimized for batch analysis of movies. • MSM analysis is done with pyTFM from Bauer et al. [2].

Homologous Recombination and DNA Intermediates Analyzed by Electron Microscopy.

Homologous recombination (HR) is a high-fidelity DNA repair pathway that uses a homologous DNA sequence as a template. Recombinase proteins are the central HR players in the three kingdoms of life. RecA/RadA/Rad51 assemble on ssDNA, generated after the processing of double-strand breaks or stalled replication forks into an active and dynamic presynaptic helical nucleofilament. Presynaptic filament formation is regulated by a series of partners of the recombinase, such as scRad52/hBRCA2 mediators or anti-recombinase proteins, to form an active machinery involved in homology search, pair-matching, and invasion within homologous sequences. During homology search, but also during strand invasion, the multiprotein complexes that form the nucleofilament induce the formation of a variety of DNA intermediate states. Here we present specific approaches to study and characterize the different DNA and DNA-protein intermediates formed during homologous recombination. The combination of powerful electron microscopy and sample preparation methods provides a better understanding of these proteins' molecular activity and their interactions.

The ionic liquid-assisted sample preparation method pTRUST allows sensitive proteome characterization of a variety of bacterial endospores to aid in the search for protein biomarkers.

Bacterial endospores are ubiquitous and are responsible for various human infections. Recently, we reported that an ionic liquid (IL)-based sample preparation method (named pTRUST) facilitated highly efficient shotgun analysis of the Bacillus subtilis spore proteome in trace samples. In this study, we evaluated the efficiency and applicability of the pTRUST technology using three different spore preparations: one purified from the closely related subspecies B. subtilis natto and two from B. licheniformis and B. cereus. We showed that the pTRUST method allowed rapid solubilization and processing of all tested spore samples prepared for highly sensitive mass spectrometry (MS) analysis. Bioinformatics analysis using the BLAST program suggested that a set of 25 proteins commonly identified between the above three species and B. subtilis spores may be universal biomarkers among various bacterial species, including 43 spore-producing bacteria associated with industrial dairy processing environments and product spoilage. In contrast, the two identified proteins, D4FV94 in B. subtilis natto and Q737A2 in B. cereus, are likely species-specific biomarkers, because their orthologs are absent or rare in all organisms. The sensitivity and applicability of pTRUST, along with the putative protein biomarkers identified in this study, will facilitate a wide spectrum of spore research for biological and clinical applications.