Sample Volume Research Articles

Description of a 'plankton filtration bias' in sequencing-based bacterial community analysis and of an Arduino microcontroller-based flowmeter device that can help to resolve it.

Diversity studies of aquatic picoplankton (bacterioplankton) communities using size-class filtration, DNA extraction, PCR and sequencing of phylogenetic markers, require a robust methodological pipeline, since biases have been demonstrated essentially at all levels, including DNA extraction, primer choice and PCR. Even different filtration volumes of the same plankton sample and, thus, different biomass loading of the filters, can distort the sequencing results. In this study, we designed an Arduino microcontroller-based flowmeter that records the decrease of initial (maximal) flowrate as proxy for increasing biomass loading and clogging of filters during plankton filtration. The device was tested using freshwater plankton of Lake Constance, and total DNA was extracted and an 16S rDNA amplicon was sequenced. We confirmed that different filtration volumes used for the same water sample affect the sequencing results. Differences were visible in alpha and beta diversities and across all taxonomic ranks. Taxa most affected were typical freshwater Actinobacteria and Bacteroidetes, increasing up to 38% and decreasing up to 29% in relative abundance, respectively. In another experiment, a lake water sample was filtered undiluted and three-fold diluted, and each filtration was stopped once the flowrate had reduced to 50% of initial flowrate, hence, at the same degree of filter clogging. The three-fold diluted sample required three-fold filtration volumes, while equivalent amounts of total DNA were extracted and differences across all taxonomic ranks were not statistically significant compared to the undiluted controls. In conclusion, this work confirms a volume/biomass-dependent bacterioplankton filtration bias for sequencing-based community analyses and provides an improved procedure for controlling biomass loading during filtrations and recovery of equivalent amounts of DNA from samples independent of the plankton density. The application of the device can also avoid the distorting of sequencing results as caused by the plankton filtration bias.

Nickel−manganese−cobalt tetragonal spinel ternary oxide nanocomposite as an effective adsorbent for dispersive solid phase micro-extraction of cadmium in food and water samples

Nickel−manganese−cobalt tetragonal spinel ternary oxide nanocomposite (NMC-TSO) was synthesized. It was utilized as an efficient sorbent for the dispersive solid phase microextraction (D-SPμE) without vortexing of cadmium. The analysis of the cadmium was carried out by FAAS. The effective analytical parameters including pH (6) contact times (no vortexing), sample volume (70 mL), eluent volume (3 mL of 2 mol L−1 HCl), linear dynamic ranges (1.07–85.7 μg L−1), and re-useability (33) on the D-SPμE efficiency were investigated. The PF, RSD% and LOD of the D-SPμE for cadmium were 23.3, ≤ 2.8% and 0.49 μg L−1, respectively. The tolerable concentrations of Ca2+, Mg2+, K+ and Na+ on Cd(II) were 50,000 mg L−1, 50,000 mg L−1, 25,000 mg L−1 and 7500 mg L−1, respectively. The method was accurated by analysis of food and water certificate reference materials (NW-TMDA-54.6 Lake water, SPS-WW1 121 Batch wastewater, 1573a Tomato Leaves and TORT-3 Lobster Hepatopancreas) and - recovery experiments. The D-SPμE-FAAS method was applied for the cadmium determination in dam water, wastewater, river water, well water, sea water, tea, cacao, nut, bitter chocolate, rice, leek, cinnamon and parsley.

EchoGrid: High-Throughput Acoustic Trapping for Enrichment of Environmental Microplastics.

The health hazards of micro- and nanoplastic contaminants in drinking water has recently emerged as an area of concern to policy makers and industry. Plastic contaminants range in size from micro- (5 mm to 1 μm) to nanoplastics (<1 μm). Microfluidics provides many tools for particle manipulation at the microscale, particularly in diagnostics and biomedicine, but has in general a limited capacity to process large volumes. Drinking water and environmental samples with low-level contamination of microplastics require processing of deciliter to liter sample volumes to achieve statistically relevant particle counts. Here, we introduce the EchoGrid, an acoustofluidics device for high throughput continuous flow particle enrichment into a robust array of particle clusters. The EchoGrid takes advantage of highly efficient particle capture through the integration of a micropatterned transducer for surface displacement-based acoustic trapping in a glass and polymer microchannel. Silica seed particles were used as anchor particles to improve capture performance at low particle concentrations and high flow rates. The device was able to maintain the silica grids at a flow rate of 50 mL/min. In terms of enrichment, the device is able to double the final pellet's microplastic concentration every 78 s for 23 μm particles and every 51 s for 10 μm particles at a flow rate of 5 mL/min. In conclusion, we demonstrate the usefulness of the EchoGrid by capturing microplastics in challenging conditions, such as large sample volumes with low microparticle concentrations, without sacrificing the potential of integration with downstream analysis for environmental monitoring.

Porosity Characteristics of Coal Seams and the Control Mechanisms of Coal Petrology in the Xishanyao Formation in the Western Part of the Southern Junggar Basin

The porosity characteristics of coal seams serve as a pivotal factor in assessing the development potential of coalbed methane (CBM) resources, significantly influencing the adsorption and permeability capabilities of coal reservoirs, as well as the accumulation, entrapment, and preservation of CBM. In this study, we focused on the coal seams of the Xishanyao Formation in the western part of the southern Junggar Basin (NW China). By leveraging the complementarity of nuclear magnetic resonance (NMR), low-temperature liquid nitrogen experiments, and high-pressure mercury intrusion porosimetry (MIP) in spatial exploration range and precision, we conducted a comprehensive analysis to achieve a fine description of porosity characteristics. Furthermore, we explored the coal petrology factors controlling the pore characteristics of the Xishanyao Formation, aiming to provide geological evidence for the selection of favorable areas and the development potential evaluation of CBM in the study area. The results indicate the following: (1) The total pore volume of the coal samples is 6.318 × 10−3 cm3/g on average, and the micropore volume accounts for a relatively high proportion (averaging 44.17%), followed by the fine pores (averaging 39.41%). The average porosity is approximately 3.87%, indicating good gas storage and connectivity of the coal seams, albeit with some heterogeneity. The coal reservoir is dominated by micropores and fine pores with diameters less than 100 nm, and the pore structure is characterized by low pore volume and high pore area. (2) The pore structure is influenced by both the coalification degree and the coal maceral. Within the range of low coalification, porosity increases with the increase in coalification degree. Building upon this, an increase in the vitrinite content promotes the development of micropores and fine pores, while an increase in the inertinite content promotes the development of meso–macropores. The clay mineral content exhibits a negative correlation with the adsorption pore volume ratio and a positive correlation with the seepage pore volume ratio.

A miniaturised semi-dynamic in-vitro model of human digestion

Reliable in-vitro digestion models that are able to successfully replicate the conditions found in the human gastrointestinal tract are key to assess the fate and efficiency of new formulations aimed for oral consumption. However, current in-vitro models either lack the capability to replicate crucial dynamics of digestion or require large volumes of sample/reagents, which can be scarce when working with nanomaterials under development. Here, we propose a miniaturised digestion system, a digestion-chip, based on incubation chambers integrated on a polymethylmethacrylate device. The digestion-chip incorporates key dynamic features of human digestion, such as gradual acidification and gradual addition of enzymes and simulated fluids in the gastric phase, and controlled gastric emptying, while maintaining low complexity and using small volumes of sample and reagents. In addition, the new approach integrates real-time automated closed-loop control of two key parameters, pH and temperature, during the two main phases of digestion (gastric and intestinal) with an accuracy down to ± 0.1 °C and ± 0.2 pH points. The experimental results demonstrate that the digestion-chip successfully replicates the gold standard static digestion INFOGEST protocol and that the semi-dynamic digestion kinetics can be reliably fitted to a first kinetic order model. These devices can be easily adapted to dynamic features in an automated, sensorised, and inexpensive platform and will enable reliable, low-cost and efficient assessment of the bioaccessibility of new and expensive drugs, bioactive ingredients or nanoengineered materials aimed for oral consumption, thereby avoiding unnecessary animal testing.

Enhancing the DNA yield intended for microbial sequencing from a low-biomass chlorinated drinking water

DNA extraction yield from drinking water distribution systems and premise plumbing is a key metric for any downstream analysis such as 16S amplicon or metagenomics sequencing. This research aimed to optimize DNA yield from low-biomass (chlorinated) reverse osmosis-produced tap water by evaluating the impact of different factors during the DNA extraction procedure. The factors examined are (1) the impact of membrane materials and their pore sizes; (2) the impact of different cell densities; and (3) an alternative method for enhancing DNA yield via incubation (no nutrient spiking). DNA from a one-liter sampling volume of RO tap water with varying bacterial cell densities was extracted with five different filter membranes (mixed ester cellulose 0.2 μm, polycarbonate 0.2 μm, polyethersulfone 0.2 and 0.1 μm, polyvinylidene fluoride 0.1 μm) for biomass filtration. Our results show that (i) smaller membrane pore size solely did not increase the DNA yield of low-biomass RO tap water; (ii) the DNA yield was proportional to the cell density and substantially dependent on the filter membrane properties (i.e., the membrane materials and their pore sizes); (iii) by using our optimized DNA extraction protocol, we found that polycarbonate filter membrane with 0.2 μm pore size markedly outperformed in terms of quantity (DNA yield) and quality (background level of 16S gene copy number) of recovered microbial DNA; and finally, (iv) for one-liter sampling volume, incubation strategy enhanced the DNA yield and enabled accurate identification of the core members (i.e., Porphyrobacter and Blastomonas as the most abundant indicator taxa) of the bacterial community in low-biomass RO tap water. Importantly, incorporating multiple controls is crucial to distinguish between contaminant/artefactual and true taxa in amplicon sequencing studies of low-biomass RO tap water.

#1094 Hemodiafiltration improves solute clearance without increasing water consumption: an educational bench model for the dialysis ward

Abstract Background and Aims Recently, the positive results of the CONVICE trial have raised the question whether the improved outcome of postdilution high volume HDF (oHDF-p) compared to high flux HD (hfHD) requires increased water consumption. From mathematical modelling it can be shown that oHDF-p always leads to higher low molecular clearance than hfHD at equal total dialysate consumption Qd_tot = Qd_dial + Qs (Qd_dial: dialysate flow through the dialyzer, Qs: substitution flow). This has also been shown in a clinical trial (Mesic et al, HDI 15, p. 522-529). However, mathematical modelling might not be intuitive for everyone, and clinical trials are always influenced by multiple confounders. To add a more intuitive way of reasoning, a simple educational bench model was designed based only on equipment easily available in a standard dialysis clinic which allows to compare dialysis efficiencies based on the urea concentrations as surrogate parameter. Since the fundamental task of dialysis is to cleanse blood from uremic solutes, the treatment modality which at equal resource utilization reduces solute concentration most is the more efficient one! Method Dialysis treatments were simulated in single pass setup using a Fresenius 5008S machine (Fig. 1). Blood inlet was simulated as a fixed urea concentration of 170 mg/dL in dialysate from a 110 L reservoir “cbi”. Treatment parameters were set such that they correspond to effective values in in-vivo treatments: As the mass transfer coefficient is always much higher in vitro with water than in vivo with blood, a small high-flux dialyzer (Fresenius FX40) was used. Blood flow Qb was set to 300 mL/min, corresponding to the in vivo blood water flow for Qb_set ∼ 350 mL/min. Alternating experiments in oHDF-p and hfHD were carried out in triplicate with Qd_tot=400 mL/min and 300 mL/min. This resulted in the settings HDF400, HD400, HDF300 and HD300 (cf. Fig. 1). Qs was set to 100 ml/min in oHDF-p. In each setting the fluid from blood and dialysate outlet were collected (recipients “cbo” and “cdo”, resp.) for 8 min. To be independent of the machine accuracy of set flow rates, sample volumes were measured by weighing, and effective flow rates of blood Qb_eff and total dialysate Qd_tot_eff were calculated. Samples were taken into monovettes from “cbi”, “cbo” and “cdo”. Urea concentrations cbi, cbo and cdo were determined by sending the samples to two commercial clinical labs who delivered consistent results. Blood and dialysate side urea clearances were calculated as KUB= Qb_eff*(cbi-cbo)/cbi and KUD=Qd_tot_eff*cdo/cbi. Results The table in Fig. 1 includes the measure values of Qb_eff and Qd_tot_eff for each setting. Blood and dialysate outlet concentrations (cbo and cdo, resp.) and calculated blood and dialysate side clearances are shown in Fig. 2. Lab results in repeated settings were highly reproducible with a SD &amp;lt; 2 mg/dL. Switching from hfHD to oHDF-p at equal Qd_tot always decreased cbo, corresponding to a higher clearance in oHDF-p. Reducing total dialysate flow from 400 to 300 ml/min caused a slight decrease in clearance in both modalities. KUB and KUD were in good agreement, showing that urea mass balance was well controlled in the experiment. Conclusion The educational bench model demonstrates the superior low molecular clearance of oHDF-p compared to hfHD at equal Qd_tot, reflected by lower urea cbo values in oHDF-p. Although the clinical outcome advantage of oHDF-p is rather related to middle molecular clearance, the model shows that the improved treatment quality of oHDF-p can be provided with no extra resource utilization compared to hfHD, and that dialysis adequacy as measured by urea clearance is even increased despite of the fact that a part of Qd_tot is deviated as substitution fluid. As moderately reducing Qd_tot in many cases only results in acceptable clearance reductions, the automated adjustment of both dialysate and substitution flow rates for oHDF-p could improve patient outcome at even reduced resource utilization.

Frequency multiplexing enables parallel multi-sample EPR

Electron paramagnetic resonance (EPR) spectroscopy stands out as a powerful analytical technique with extensive applications in the fields of biology, chemistry, physics, and material sciences. It proves invaluable for investigating the molecular structure and reaction mechanisms of substances containing unpaired electrons, such as metal complexes, organic and inorganic radicals, and intermediate states in chemical reactions. However, despite their remarkable capabilities, EPR systems face significant limitations in terms of sample throughput, as current commercial systems only target the analysis of one sample at a time. Here we introduce a novel scheme for conducting ultra-high frequency continuous-wave EPR (CW EPR) targeting the EPR spectroscopy of multiple microliter volume samples in parallel. Our proof-of-principle prototype involves two decoupled detection cells equipped with high qualty factor Q=104\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$Q=104$$\\end{document} solenoidal coils tuned to 488 and 589 MHz, ensuring a significant frequency gap for effective radio frequency (RF) decoupling between the channels. To further enhance electromagnetic decoupling, an orthogonal alignment of the coils was adopted. The paper further presents an innovative radiofrequency circuit concept that utilizes a single physical RF channel to simultaneously conduct parallel EPR on up to eight cells. Parallel EPR experiments on two BDPA samples, each with a sample volume of 18.3 μL, registered signal-to-noise ratios of 255 and 252 for the two EPR measurement cells, with no observable coupling. The showcased prototype, built using cost-effective commercially available fabrication technology, is readily scalable and represents an initial step with promising potential for advancing sample screening with high-throughput parallel EPR.

Characteristics of lipid metabolism after treatment of colon cancer mice with American ginseng vesicles.

Lipid molecules are present in tumours and play an important role in the anti-inflammatory response as well as in antiviral protection. Changes in the type and location of lipids in the intestine following exposure to environmental stressors play an important role in several disorders, including ulcerative colitis (UC), inflammatory bowel disease (IBD), and colorectal cancer. The aim of this work is to provide a new theoretical basis for tumour initiation and development by accurately measuring the spatial distribution of lipids and metabolites in intestinal tissue. Spatial metabolomics allows the detection of samples with minimal sample volume by label-free imaging of complex samples in their original state. The distribution of lipid molecules in tumours has not been reported, although the distribution of lipid molecules in intestinal tissue has been reported in the literature. The range of lipid profiles in colon cancer mouse tumour tissue was compiled using a spatial metabolomics: lipid extraction method. The changes in lipid distribution in two regions after oral administration of American Ginseng (Panax quinquefolius L.) vesicles were also compared. Tumour tissue samples were extracted with 80% methanol-20% formic acid in water. The resulting spatial metabolic profile allowed the identification of seven lipid classes in mouse tumours. The distribution of fibre tissue cells was 23.2% higher than tumour tissue cells, with the exception of the fatty acid (FA) species.

Exploring toroidal anvil profiles for larger sample volumes above 4 Mbar.

With the advent of toroidal and double-stage diamond anvil cells (DACs), pressures between 4 and 10 Mbar can be achieved under static compression, however, the ability to explore diverse sample assemblies is limited on these micron-scale anvils. Adapting the toroidal DAC to support larger sample volumes offers expanded capabilities in physics, chemistry, and planetary science: including, characterizing materials in soft pressure media to multi-megabar pressures, synthesizing novel phases, and probing planetary assemblages at the interior pressures and temperatures of super-Earths and sub-Neptunes. Here we have continued the exploration of larger toroidal DAC profiles by iteratively testing various torus and shoulder depths with central culet diameters in the 30-50µm range. We present a 30µm culet profile that reached a maximum pressure of 414(1) GPa based on a Pt scale. The 300K equations of state fit to our P-V data collected on gold and rhenium are compatible with extrapolated hydrostatic equations of state within 1% up to 4 Mbar. This work validates the performance of these large-culet toroidal anvils to > 4 Mbar and provides a promising foundation to develop toroidal DACs for diverse sample loading and laser heating.

A new methodology for a rapid and high-throughput comparison of molecular profiles and biological activity of phytoextracts.

To robustly discover and explore phytocompounds, it is necessary to evaluate the interrelationships between the plant species, plant tissue, and the extraction process on the extract composition and to predict its cytotoxicity. The present work evaluated how Fourier Transform InfraRed spectroscopy can acquire the molecular profile of aqueous and ethanol-based extracts obtained from leaves, seeds, and flowers of Cynara Cardunculus, and ethanol-based extracts from Matricaria chamomilla flowers, as well the impact of these extracts on the viability of mammalian cells. The extract molecular profile enabled to predict the extraction yield, and how the plant species, plant tissue, and extraction process affected the extract's relative composition. The molecular profile obtained from the culture media of cells exposed to extracts enabled to capture its impact on cells metabolism, at a higher sensitivity than the conventional assay used to determine the cell viability. Furthermore, it was possible to detect specific impacts on the cell's metabolism according to plant species, plant tissue, and extraction process. Since spectra were acquired on small volumes of samples (25 µL), after a simple dehydration step, and based on a plate with 96 wells, the method can be applied in a rapid, simple, high-throughput, and economic mode, consequently promoting the discovery of phytocompounds.

Multiple-scattering effects on single-wavelength lidar sounding of multi-layered clouds

Abstract. We performed Monte Carlo simulations of single-wavelength lidar signals from multi-layered clouds with special attention focused on the multiple-scattering (MS) effect in regions of the cloud-free molecular atmosphere (i.e. between layers or outside a cloud system). Despite the fact that the strength of lidar signals from the molecular atmosphere is much lower compared to the in-cloud intervals, studies of MS effects in such regions are of interest from scientific and practical points of view. The MS effect on lidar signals always decreases with the increasing distance from the cloud far edge. The decrease is the direct consequence of the fact that the forward peak of particle phase functions is much larger than the receiver field of view (RFOV). Therefore, the photons scattered within the forward peak escape the sampling volume formed by the RFOV (i.e. the escape effect). We demonstrated that the escape effect is an inherent part of MS properties within the free atmosphere beyond the cloud far edge. In the cases of the ground-based lidar, the MS contribution is lower than 5 % within the regions of the cloud-free molecular atmosphere with a distance from the cloud far edge of about 1 km or higher. In the cases of the space-borne lidar, the rate of decrease of the MS contribution is so slow that the threshold of 5 % can hardly be reached. In addition, the effect of non-uniform beam filling is extremely strong. Therefore, practitioners should employ, with proper precautions, lidar data from regions below the cloud base when treating data of a space-borne lidar. In the case of two-layered cloud, the distance of 1 km is sufficiently large so that the scattered photons emerging from the first layer do not affect signals from the second layer when we are dealing with the ground-based lidar. In contrast, signals from the near edge of the second cloud layer are severely affected by the photons emerging from the first layer in the case of a space-borne lidar. We evaluated the Eloranta model (EM) in extreme conditions and showed its good performance in the cases of ground-based and space-borne lidars. At the same time, we revealed the shortcoming that can affect practical applications of the EM. Namely, values of the key parameters – i.e. the ratios of phase functions in the backscatter direction for the nth-order-scattered photon and a singly scattered photon – depend not only on the particle phase function but also on the distance from a lidar to the cloud and the receiver field of view. Those ratios vary within a quite large range, and the MS contribution to lidar signals can be largely overestimated or underestimated if erroneous values of the ratios are assigned to the EM.

Hybrid Hydrogel-Magnet Actuated Capsule for Automatic Gut Microbiome Sampling.

Non-invasive, pill-sized capsules can provide intestinal fluid sampling to easily retrieve site-specific gut microbiome samples for studies in nutrition and chronic diseases. However, capsules with both automatic sampling and active locomotion are uncommon due to limited onboard space. This paper presents a novel hybrid hydrogel-magnet actuated capsule featuring: i) pH-responsive hydrogels that will automatically trigger fluid sampling at an environmental pH of > 6 and ii) active locomotion by an external rotating magnetic field. Two capsule designs were fabricated (Design A: 31 μL sampling volume with dimensions 8 mm × 19 mm, Design B: 41 μL sampling volume with dimensions 8 mm × 21 mm). They were immersed in simulated gastric (pH = 1.2) and simulated intestinal fluid (pH = 6.8) to test for automatic intestinal fluid sampling. An external rotating magnetic field was applied to test for active locomotion. Finally, seal tests were performed to demonstrate sample contamination mitigation. Preliminary experiments showed that sampling occurred quickly and automatically in simulated intestinal fluid at 6 - 15 hours, active locomotion via rotation, rolling, and tumbling were possible at magnetic field magnitudes < 10 mT, oil piston seals were better at mitigating sample contamination than water piston seals, and minimum o-ring seal pressures limits of 1.95 and 1.69 kPa for Design A and B respectively were sufficient against intra-abdominal pressures. This work presents the ability to impart capsule multi-functionality in a compact manner without onboard electronics or external triggering for sampling.

Molecular fingerprinting of biological nanoparticles with a label-free optofluidic platform

Label-free detection of multiple analytes in a high-throughput fashion has been one of the long-sought goals in biosensing applications. Yet, for all-optical approaches, interfacing state-of-the-art label-free techniques with microfluidics tools that can process small volumes of sample with high throughput, and with surface chemistry that grants analyte specificity, poses a critical challenge to date. Here, we introduce an optofluidic platform that brings together state-of-the-art digital holography with PDMS microfluidics by using supported lipid bilayers as a surface chemistry building block to integrate both technologies. Specifically, this platform fingerprints heterogeneous biological nanoparticle populations via a multiplexed label-free immunoaffinity assay with single particle sensitivity. First, we characterise the robustness and performance of the platform, and then apply it to profile four distinct ovarian cell-derived extracellular vesicle populations over a panel of surface protein biomarkers, thus developing a unique biomarker fingerprint for each cell line. We foresee that our approach will find many applications where routine and multiplexed characterisation of biological nanoparticles are required.

Double-Sided Tape in Microfluidics: A Cost-Effective Method in Device Fabrication.

The demand for easy-to-use, affordable, accessible, and reliable technology is increasing in biological, chemical, and medical research. Microfluidic devices have the potential to meet these standards by offering cost-effective, highly sensitive, and highly specific diagnostic tests with rapid performance and minimal sample volumes. Traditional microfluidic device fabrication methods, such as photolithography and soft lithography, are time-consuming and require specialized equipment and expertise, making them costly and less accessible to researchers and clinicians and limiting the applicability and potential of microfluidic devices. To address this, researchers have turned to using new low-cost materials, such as double-sided tape for microfluidic device fabrication, which offers simple and low-cost processes. The innovation of low-cost and easy-to-make microfluidic devices improves the potential for more devices to be transitioned from laboratories to commercialized products found in stores, offices, and homes. This review serves as a comprehensive summary of the growing interest in and use of double-sided tape-based microfluidic devices in the last 20 years. It discusses the advantages of using double-sided tape, the fabrication techniques used to create and bond microfluidic devices, and the limitations of this approach in certain applications.

A simple and highly sensitive LC–MS workflow for characterization and quantification of ADC cleavable payloads

Antibody–drug conjugates (ADC) payloads are cleavable drugs that act as the warhead to exert an ADC’s cytotoxic effects on cancer cells intracellularly. A simple and highly sensitive workflow is developed and validated for the simultaneous quantification of six ADC payloads, namely SN-38, MTX, DXd, MMAE, MMAF and Calicheamicin (CM). The workflow consists of a short and simple sample extraction using a methanol-ethanol mixture, followed by a fast liquid chromatography tandem mass spectrometry (LC–MS/MS) analysis. The results showed that well-validated linear response ranges of 0.4–100 nM for SN38, MTX and DXd, 0.04–100 nM for MMAE and MMAF, 0.4–1000 nM for CM were achieved in mouse serum. Recoveries for all six payloads at three different concentrations (low, medium and high) were more than 85%. An ultra-low sample volume of only 5 µL of serum is required due to the high sensitivity of the method. This validated method was successfully applied to a pharmacokinetic study to quantify MMAE in mouse serum samples.

Synthesis of nano magnetic chitosan modified with acrylamide schiff’s base for simultaneous removal of Cu(II) and Co(II) from waste water samples

ABSTRACT A magnetic nano sorbent based on the functionalization of magnetic chitosan with acrylamide was synthesized for Cu(II) and Co(II) removal from wastewater samples. To characterize the magnetic nano sorbent, they have been applied from Fourier transform infrared spectroscopic analysis (FTIR), X-ray diffraction (XRD) and scanning electron microscopy (SEM) techniques. The effective parameters of Cu(II) and Co(II) adsorption on magnetic nano sorbent such as pH of the solution, adsorbent dose, contact time and sample volume, were optimized. Also, the effective parameters on Cu(II) and Co(II) desorption conditions (type and volume of elution solvent) were optimized. The isotherms and kinetics models for the adsorption of Cu(II) and Co(II) on the magnetic nano sorbent were studied. The data were well-fitted with the Langmuir isotherm and pseudo-second-order model. The maximum adsorption capacities for Cu(II) and Co(II) were 165.4 and 159.9 mg g-1, respectively. Also, the thermodynamic parameters were studied. Application of magnetic nano sorbent for 4 times of use was acceptable. For the real samples, the percentage of recovery was higher than 97% and RSD (%) was between 1.4% and 2.7%. Finally, the magnetic nano sorbent exhibited high performance in removing the Cu(II) and Co(II) from wastewater samples.

Label-free and label-based electrochemical detection of disease biomarker proteins

Introduction: Biosensors, analytical devices integrating biological sensing elements with physicochemical transducers, have gained prominence as rapid and convenient tools for monitoring human health status using biochemical analytes. Due to its cost-effectiveness, simplicity, portability, and user-friendliness, electrochemical detection has emerged as a widely adopted method in biosensor applications. Crucially, biosensors enable early disease diagnosis by detecting protein biomarkers associated with various conditions. These biomarkers offer an objective indication of medical conditions that can be accurately observed from outside the patient. Method: This review comprehensively documents both label-free and labelled detection methods in electrochemical biosensor techniques. Label-free detection mechanisms elicit response signals upon analyte molecule binding to the sensor surface, while labelled detection employs molecular labels such as enzymes, nanoparticles, and fluorescent tags. Conclusion: The selection between label-free and labelled detection methods depends on various factors, including the biomolecular compound used, analyte type and biological binding site, biosensor design, sample volume, operational costs, analysis time, and desired detection limit. Focusing on the past six years, this review highlights the application of label-free and labelled electrochemical biosensors for detecting protein biomarkers of diseases.

Top-Down Proteomics Analysis of Picogram-Level Complex Samples Using Spray-Capillary-Based Capillary Electrophoresis-Mass Spectrometry.

Proteomics analysis of mass-limited samples has become increasingly important for understanding biological systems in physiologically relevant contexts such as patient samples, multicellular organoids, spheroids, and single cells. However, relatively low sensitivity in top-down proteomics methods makes their application to mass-limited samples challenging. Capillary electrophoresis (CE) has emerged as an ideal separation method for mass-limited samples due to its high separation resolution, ultralow detection limit, and minimal sample volume requirements. Recently, we developed "spray-capillary", an electrospray ionization (ESI)-assisted device, that is capable of quantitative ultralow-volume sampling (e.g., pL-nL level). Here, we developed a spray-capillary-CE-MS platform for ultrasensitive top-down proteomics analysis of intact proteins in mass-limited complex biological samples. Specifically, to improve the sensitivity of the spray-capillary platform, we incorporated a polyethylenimine (PEI)-coated capillary and optimized the spray-capillary inner diameter. Under optimized conditions, we successfully detected over 200 proteoforms from 50 pg of E. coli lysate. To our knowledge, the spray-capillary CE-MS platform developed here represents one of the most sensitive detection methods for top-down proteomics. Furthermore, in a proof-of-principle experiment, we detected 261 ± 65 and 174 ± 45 intact proteoforms from fewer than 50 HeLa and OVCAR-8 cells, respectively, by coupling nanodroplet-based sample preparation with our optimized CE-MS platform. Overall, our results demonstrate the capability of the modified spray-capillary CE-MS platform to perform top-down proteomics analysis on picogram amounts of samples. This advancement presents the possibility of meaningful top-down proteomics analysis of mass-limited samples down to the level of single mammalian cells.

Persistence and risk factors of occult hepatitis B virus infections among antiretroviral therapy-naïve people living with HIV in Botswana.

This study aimed to determine the kinetics of occult hepatitis B virus infections (OBI) among people with HIV (PWH). The study used archived plasma samples from longitudinal HIV natural history studies. We identified new OBI cases and assessed risk factors for OBI using Cox proportional hazards regression analysis. At baseline, 8 of 382 [(2.1%) (95% CI: 1.06-4.1)] samples tested positive for hepatitis B surface antigen (HBsAg+). Of the 374 HBsAg-negative samples, 76 had sufficient sample volume for HBV DNA screening. OBI positivity (OBI+) at baseline was reported in 11 of 76 [14.7 95% CI (8.3-24.1)] HBsAg-negative (HBsAg-) participants. Baseline HBsAg-negative samples with sufficient follow-up samples (n = 90) were used for analysis of newly identified OBI cases. Participants contributed 129.74 person-years to the study and were followed for a median of 1.02 years (IQR: 1.00-2.00). Cumulatively, there were 34 newly identified OBI cases from the 90 participants, at the rate of 26.2/100 person-years (95% CI: 18.7-36.7). Newly identified OBI cases were more common among men than women (61.1% vs. 31.9%) and among participants with CD4+ T-cell counts ≤450 cells/mL (p-value = 0.02). Most of the newly identified OBI cases [55.9% (19/34)] were possible reactivations as they were previously HBV core antibody positive. There was a high rate of newly identified OBI among young PWH in Botswana, especially in men and in participants with lower CD4+ T-cell counts. OBI screening in PWH should be considered because of the risk of transmission, possible reactivation, and risk factors for the development of chronic liver disease, including hepatocellular carcinoma.