Limit Of Dynamic Range Research Articles

Ultrasensitive bioassaying of HER-2 protein for diagnosis of breast cancer using reduced graphene oxide/chitosan as nanobiocompatible platform

BackgroundIn this label-free bioassay, an electrochemiluminescence (ECL) immunosensor was developed for the quantification of breast cancer using HER-2 protein as a metastatic biomarker.MethodFor this purpose, the ECL emitter, [Ru(bpy)3]2+, was embedded into biocompatible chitosan (CS) polymer. The prepared bio-composite offered high ECL reading due to the depletion of human epidermal growth factor receptor 2 (HER-2) protein. Reduced graphene oxide (rGO) was used as substrate to increase signal stability and achieve greater sensitivity. For this, rGO was initially placed electrochemically on the glassy carbon electrode (GCE) surface by cyclic voltammetry (CV) technique. Next, the prepared CS/[Ru(bpy)3]2+ biopolymer solution was coated on a drop of the modified electrode such that the amine groups of CS and the carboxylic groups of rGO could covalently interact. Using EDC/NHS chemistry, monoclonal antibodies (Abs) of HER-2 were linked to CS/[Ru(bpy)3]2+/rGO/GCE via amide bonds between the carboxylic groups of Ab molecules and amine groups of CS. The electrochemical behavior of the electrode was studied using different electrochemical techniques such as electrochemical impedance spectroscopy (EIS), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) and also ECL tests.ResultsAfter passing all optimization steps, the lower limit of detection (LLOQ) and linear dynamic range (LDR) of HER-2 protein were practically obtained as 1 fM and 1 fM to 1 nM, individually. Importantly, the within and between laboratory precisions were performed and the suitable relative standard deviations (RSDs) were recorded as 3.1 and 3.5%, respectively.ConclusionsAs a proof of concept, the designed immunosensor was desirably applied for the quantification of HER-2 protein in breast cancer suffering patients. As a result, the designed ECL-based immunosensor has the capability of being used as a conventional test method in biomedical laboratories for early detection of HER-2 protein in biological fluids.Graphic

Nanoporous hydrogel for direct digital nucleic acid amplification in untreated complex matrices for single bacteria counting

Direct quantification of pathogens in unprocessed complex samples remain challenging due to the severe inhibition of nucleic acid amplification. In this work, we report a nanoporous polyethylene glycol hydrogel with self-cleaning capacity for direct amplification of nucleic acid in complex matrices (human whole blood, animal blood, milky tea, humic acid, and surfactants) without any sample pretreatment or DNA extraction. During isothermal amplification inside the hydrogel, the inhibitors in the assay will be adsorbed and removed by the surrounding nanostructured polymers, and nucleic acid amplification was proceeding successfully, resulting in a series of bright dots for single bacteria counting. Thus, the loop-mediated isothermal amplifications (LAMP) performed inside hydrogel demonstrated a high level of resistance to inhibition in various complex matrices. The underlying anti-inhibition mechanism was also investigated. Digital quantification of Escherichia coli, Salmonella typhi and Listeria monocytogenes in whole blood were achieved within 20 min, with wide dynamic range, high specificity and low detection limit down to single bacterium. Visual counting via naked eye was also successfully established with the help of a conventional LED flashlight. We believe the developed hydrogel nanofluidic system has an enormous potential for on-site direct analysis of complex, crude, and unprocessed samples in clinical, food, agricultural, and environmental fields.

Point-of-Care Pathogen Testing Using Photonic Crystals and Machine Vision for Diagnosis of Urinary Tract Infections.

Urinary tract infections (UTIs) caused by bacterial invasion can lead to life-threatening complications, posing a significant health threat to more than 150 million people worldwide. As a result, there is need for accurate and rapid diagnosis of UTIs to enable more effective treatment. Described here is an intelligent diagnostic system constructed for bacterial detection using an immunobiosensor, signal-amplification biochip, and image processing algorithm based on machine vision. This prototype can quickly detect bacteria by collection of enhanced luminescence enabled by the photonic crystals integrated into the biochip. By use of a machine vision algorithm, the very small luminescence signals are analyzed to provide a low detection limit and wide dynamic range. This sensor system can offer an affordable, accessible, and user-friendly digital diagnostic solution, possibly suitable for wearable technology, that could improve treatment of this challenging disease.

An ultra-sensitive electrochemiluminescence probe based on ternary nanocomposite and boron nitride quantum dots for detection of diazinon.

Anew enzyme-free electrochemiluminescence (ECL) pesticide sensor was fabricated based on ternary nanocomposite of ruthenium nanoparticles/silver nanoparticles/graphene oxide on the surface of glassy carbon electrode for ultratrace determination of diazinon. Due to some drawbacks of enzyme-based sensors such as enzyme instability at elevatedtemperature, humidity, changes of pH, and high price of the enzyme, the use ofenzyme was omitted in the construction of the developed sensor. The silver nanoparticles with good electrocatalytic proficiency as a signal improving agent and tris(2,2bipyridine) ruthenium(II) as a popular luminophore were uniformly deposited on the surface of the prepared graphene oxide/GC electrode at nanoscale. Boron nitride quantum dots as an efficient co-reactant created the superior efficiency in amplifying the ECL intensity of the ruthenium-based ECL system. The prepared electrode was utilized for the detection of diazinon via the robust ECL method. For the present sensor, a wide linear dynamic range and low detection limit were achieved (3.0 × 10-15 to 6.5 × 10-9M and 9.5 × 10-16M, respectively). The obtained results confirmed the fabrication of the robust ECL probe, which is characterized by the cooperative effect of silver nanoparticles and the attached luminophore species. The main advantage of the presented sensor was that the samples could be diluted so that the effect of the interference species was negligible. Due to excellent properties toward accurate determination of diazinon, the ECL sensor as a new practical platform was applied for quantitative detection of diazinon in some real samples.

In situ growth of copper-based metal–organic framework nanoarrays on copper wire for solid-phase microextraction of polycyclic aromatic hydrocarbons

The present study deals with metal–organic framework substances in three-dimensional oriented design vital for using them in a new type of the fiber coating for solid-phase micro-extraction of polycyclic aromatic hydrocarbons. Cu(OH)2 nano-rod arrays were utilized as a sacrificial template to develop a coordination replication approach to make Cu-based metal–organic frameworks into three-dimensional hierarchical nano-array architectures. The Cu(OH)2 nano-rod arrays served as Cu source coordinating with organic ligand to create MOF crystals and the three-dimensional substrate for supporting the Cu-based metal–organic framework growth. By this strategy, novel possibilities are made to construct immobilized metal–organic framework-based three dimensional nano-arrays for multipurpose usages. Optimum circumstances are a desorption temperature of 260 °C, an extraction temperature of 85 °C, and a salt concentration of 20% w/v. The limit of detection and dynamic linear range are within the range of 2.5–3.4 ngL-1 and 0.01–100 µgL-1, respectively. One fiber’s repeatability (n = 5) is between 5.1 and 8.4%, stated as relative standard deviation. The technique possesses the benefits including simple use, and the fiber’s quick thermal stability, as well as high relative recovery (in comparison with usual approaches), are further attractive properties.

Recent Advances in Nanotechnology-Based Biosensors Development for Detection of Arsenic, Lead, Mercury, and Cadmium.

Heavy metals cause considerable environmental pollution due to their extent and non-degradability in the environment. Analysis and trace levels of arsenic, lead, mercury, and cadmium as the most toxic heavy metals show that they can cause various hazards in humans’ health. To achieve rapid, high-sensitivity methods for analyzing ultra-trace amounts of heavy metals in different environmental and biological samples, novel biosensors have been designed with the participation of strategies applied in nanotechnology. This review attempted to investigate the novel, sensitive, efficient, cost-benefit, point of care, and user-friendly biosensors designed to detect these heavy metals based on functional mechanisms. The study’s search strategies included examining the primary databases from 2015 onwards and various keywords focusing on heavy metal biosensors’ performance and toxicity mechanisms. The use of aptamers and whole cells as two important bio-functional nanomaterials is remarkable in heavy metal diagnostic biosensors’ bioreceptor design. The application of hybridized nanomaterials containing a specific physicochemical function in the presence of a suitable transducer can improve the sensing performance to achieve an integrated detection system. Our study showed that in addition to both labeled and label-free detection strategies, a wide range of nanoparticles and nanocomposites were used to modify the biosensor surface platform in the detection of heavy metals. The detection limit and linear dynamic range as an essential characteristic of superior biosensors for the primary toxic metals are studied. Furthermore, the perspectives and challenges facing the design of heavy metal biosensors are outlined. The development of novel biosensors and the application of nanotechnology, especially in real samples, face challenges such as the capability to simultaneously detect multiple heavy metals, the interference process in complex matrices, the efficiency and stability of nanomaterials implemented in various laboratory conditions.

A highly sensitive electrochemiluminescence cytosensor for detection of SKBR-3 cells as metastatic breast cancer cell line: A constructive phase in early and precise diagnosis

Ultrasensitive monitoring of cancer cells, especially metastatic ones, has a great interest in human medicine. Despite the early diagnosis of diseases, there is an essential need for any prediction in the severity of side effects for therapeutic outcomes like metastasis. Therefore, the inhibition of cancer cells metastasis to other organs is of utmost importance for cancer suffering patients. In this regard, we developed an electrochemiluminescence (ECL)-based cytosensor for the quantification of metastatic breast cancer cells, namely SKBR-3. Silica-based mesoporous materials have a great potential for application in ECL biosensors due to their high loading capacity and mechanical strength. Herein, a silica-based electrode was prepared via in situ electrosyntheses of mesoporous silica as an environmentally friendly approach. In this protocol, luminol (as luminophore) was combined with chitosan (as attachment biomolecule) to produce a stable lumino-composite film on the electrode surface. At the optimum experimental conditions, the lower limit of quantitation (LLOQ) and linear dynamic range (LDR) were obtained as 20 cells/mL and 20 to 2000 cells/mL, individually. The specificity was desirably examined in the presence of other breast cancer cell lines such as MCF-7 and MDA-MB-231, as a model of early-stage and invasive phases of breast cancer cells. The repeatability was successfully examined for five repetitive measurements and the acceptable relative standard deviation (RSD) was calculated as about 1.6% for 500 cells/mL. As a proof of concept, the presented cytosensor has a high ability to use in clinical laboratories for the detection and separation of metastatic cells via the combination with microfluidic systems.

Rhodamine-based fluorescent polyacrylic nanoparticles: A highly selective and sensitive chemosensor for Fe (II) and Fe (III) cations in water

Fe2+ and specially Fe3+ are important in our daily life and their deficiency or excess amounts may cause serious disorders and infectious diseases. Most of their existing fluorescent sensors consist of small molecules and may operate in organic media with prolonged response time and inadequate detection limit which restrict their wide-spread applications. Fluorescent poly(methyl methacrylate-co-glycidyl methacrylate) nanoparticles bearing Rhodamine B ethylenediamine acrylate (RhAcL-3) were prepared through semi-continues emulsion polymerization. Chemical structure of the prepared RhAcL-3 polyacrylic nanoparticles with smooth spherical morphology, average particle size of 46 nm and narrow size distribution (polydispersity index of 0.09) were completely characterized by FTIR, 1HNMR, UV-Vis, SEM and DLS techniques. Water dispersion of RhAcL-3 nanoparticles demonstrated an immediate and apparent color change together with enhancement in fluorescence emission at 590 nm after addition of Fe2+ and Fe3+ ions up to 2 and 2.2-folds, respectively. This was attributed to chelation of these ions to spirolactam moiety in Rhodamine B to consequate open-ring amide form. It was found that the presence of some other metal cations had no interference on the above affinity and sensing role. Linear dynamic detection range (4–320 µM) and detection limit of RhAcL-3 sensor toward Fe2+ and Fe3+ were estimated 2.63 and 2.5 µM, respectively. The results suggest that these prepared nanoparticles could be utilized as colorimetric and fluorescent sensor with instant responsivity, wide linear detection range, high sensitivity and low detection limit for Fe2+ and Fe3+ ions in future advances and applications.

A multiplexed immuno-sensor for on-line and automated monitoring of tissue culture protein biomarkers

Frequent on-line and automated monitoring of multiple protein biomarkers level secreted in the culture media during tissue growth is essential for the successful development of Tissue Engineering and Regenerative Medicine (TERM) products. Here, we present a low-cost, rapid, reliable, and integrable anion-exchange membrane-(AEM) based multiplexed sensing platform for this application. Unlike the gold-standard manual ELISA test, incubation/wash steps are optimized for each target and precisely metered in microfluidic chips to enhance selectivity. Unlike optical detection and unreliable visual detection for the ELISA test, which require standardization for every usage, the AEM ion current signal also offers robustness, endowed by the pH and ionic strength control capability of the ion-selective membrane, such that a universal standard curve can be used to calibrate all runs. The electrical signal is enhanced by highly charged silica nanoparticle reporters, which also act as hydrodynamic shear amplifiers to enhance selectivity during wash. This AEM-based sensing platform is tested with vascular protein biomarkers, Endothelin-1 (ET-1), Angiogenin (ANG) and Placental Growth Factor (PlGF). The limit of detection and three-decade dynamic range are comparable to ELISA assay but with a significantly reduced assay time of 1 h vs 7 h, due to the elimination of calibration and blocking steps. Optimized protocol for each target renders the detection highly reliable with more than 98% confidence. The multiplexed detection capability of the platform is also demonstrated by simultaneous detection of ET-1, ANG and PlGF in 40 μl of the vascular endothelial cell culture supernatants using three-membrane AEM sensor and the performance is validated against ELISA.

Label-Free Electrochemical Aptasensors Based on Photoresist Derived Carbon for Cancer Biomarker Detection

Cancer biomarkers are substances found in the human fluid, tissue, and bone marrow that can indicate the presence of cancer diseases in the human body. In the past decades, various cancer biomarkers have been discovered for early detection of cancers and risk assessment of reoccurring cancers. Among the discovered cancer biomarkers, platelet-derived growth factor-BB (PDGF-BB) is an essential biomarker for early detection of cancer diseases. This biomarker plays a significant role in the development of multiple solid malignant tumors (e.g., breast, brain, pancreatic, prostate, and ovarian) and lymphatic metastasis. A feasible means for early detection of cancers is developing point-of-care biosensors based on synthetic DNA or RNA bio-recognizers (i.e., aptamers), which are referred to as aptasensors. Various advanced technologies have been investigated for developing cancer biomarker aptasensors, including optical, piezoelectric, and electrochemical based techniques. Among the developed technologies, label-free electrochemical aptasensors are highly potent means for this purpose since they can reduce the cost and complications of sample preparation, and the electrochemical cells can be efficiently miniaturized and integrated with available lab-on-chips and MEMS technologies. However, the sensing performance of the label-free electrochemical cancer aptasensors, including the limit of detection, selectivity, and dynamic range requires further enhancements to be at similar levels to benchtop testing. In the light of the importance of PDGF-BB detection and demanded enhancements for label-free electrochemical aptasensors, we have developed label-free electrochemical aptasensors based on photoresist derived carbon microelectrodes. The active electrode of this aptasensor was synthesized via well-established C-MEMS (carbon microelectromechanical systems) fabrication technology. C-MEMS platforms have distinguishing features such as low background capacitance, high stability when they exposed to different physical/chemical treatments, biocompatibility, and good electrical conductivity. Furthermore, the surface of C-MEMS can be modified effectively via electrochemical processes as well as depositing nanomaterials. The applied C-MEMS fabrication in this study includes photopatterning of SU-8 25 negative photoresist-based microelectrodes and carbonization of the developed microelectrodes at high temperatures and oxygen-free tube furnaces. The carbonized microelectrodes were functionalized utilizing oxygen plasma oxidation pretreatment to introduce carboxyl groups to the surfaces of the carbon microelectrodes. The PDGF-BB affinity aptamers were covalently immobilized via amid binding of amino-tag terminated aptamers and carboxyl groups covered carbon surfaces. The Fourier-transform infrared spectroscopy (FTIR) confirmed the successful carboxyl group functionalization of the C-MEMS microelectrodes and covalent immobilization of affinity aptamers via amide biding. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used for characterizing the C-MEMS based aptasensors in different stages of development and sensing performances. The turn-on sensing strategy was deployed via measuring the charge transfer resistance (RCT) from EIS Nyquist plots, which yielded to wide sensing linear range of 0.005 – 50 nM with a high sensitivity of 14.82 × 103 Ω.(Log (M))-1 and low limit of detection of 1.9 pM (S/N=3). The turn-off sensing strategy was applied via measuring capacitance from CV curves, which conceded to wide linear response range of 0.01 – 50 nM with a high sensitivity of 29.97 mF.cm-2.(Log (M))-1 and low limit of detection of 7 pM (S/N=3) toward the PDGF-BB. The developed label-free electrochemical aptasensor exhibited good selectivity, stability, and repeatability, which is highly promising for future lab-on-chip and point-of-care cancer diagnosis technologies.

Electrochemical Detection of Multianalyte Biomarkers in Wound Healing Efficacy.

The targeted diagnosis and effective treatments of chronic skin wounds remain a healthcare burden, requiring the development of sensors for real-time monitoring of wound healing activity. Herein, we describe an adaptable method for the fabrication of carbon ultramicroelectrode arrays (CUAs) on flexible substrates with the goal to utilize this sensor as a wearable device to monitor chronic wounds. As a proof-of-concept study, we demonstrate the electrochemical detection of three electroactive analytes as biomarkers for wound healing state in simulated wound media on flexible CUAs. Notably, to follow pathogenic responses, we characterize analytical figures of merit for identification and monitoring of bacterial warfare toxin pyocyanin (PYO) secreted by the opportunistic human pathogen Pseudomonas aeruginosa. We also demonstrate the detection of uric acid (UA) and nitric oxide (NO•), which are signaling molecules indicative of wound healing and immune responses, respectively. The electrochemically determined limit of detection (LOD) and linear dynamic range (LDR) for PYO, UA, and NO• fall within the clinically relevant concentrations. Additionally, we demonstrate the successful use of flexible CUAs for quantitative, electrochemical detection of PYO from P. aeruginosa strains and cellular NO• from immune cells in the wound matrix. Moreover, we present an electrochemical examination of the interaction between PYO and NO•, providing insight into pathogen-host responses. Finally, the effects of the antimicrobial agent, silver (Ag+), on P. aeruginosa PYO production rates are investigated on flexible CUAs. Our electrochemical results show that the addition of Ag+ to P. aeruginosa in wound simulant decreases PYO secretion rates.

Improved Sensitivity of Ultralow Flow LC–MS-Based Proteomic Profiling of Limited Samples Using Monolithic Capillary Columns and FAIMS Technology

In this work, we pioneered a combination of ultralow flow (ULF) high-efficiency ultranarrow bore monolithic LC columns coupled to MS via a high-field asymmetric waveform ion mobility spectrometry (FAIMS) interface to evaluate the potential applicability for high sensitivity, robust, and reproducible proteomic profiling of low nanogram-level complex biological samples. As a result, ULF LC-FAIMS-MS brought unprecedented sensitivity levels and high reproducibility in bottom-up proteomic profiling. In addition, FAIMS improved the dynamic range, signal-to-noise ratios, and detection limits in ULF LC-MS-based measurements by significantly reducing chemical noise in comparison to the conventional nanoESI interface used with the same ULF LC-MS setup. Two, three, or four compensation voltages separated by at least 15 V were tested within a single LC-MS run using the FAIMS interface. The optimized ULF LC-ESI-FAIMS-MS/MS conditions resulted in identification of 2,348 ± 42 protein groups, 10,062 ± 285 peptide groups, and 15,734 ± 350 peptide-spectrum matches for 1 ng of a HeLa digest, using a 1 h gradient at the flow rate of 12 nL/min, which represents an increase by 38%, 91%, and 131% in respective identifications, as compared to the control experiment (without FAIMS). To evaluate the practical utility of the ULF LC-ESI-FAIMS-MS platform in proteomic profiling of limited samples, approximately 100, 1,000, and 10,000 U937 myeloid leukemia cells were processed, and a one-tenth of each sample was analyzed. Using the optimized conditions, we were able to reliably identify 251 ± 54, 1,135 ± 80, and 2,234 ± 25 protein groups from injected aliquots corresponding to ∼10, 100, and 1,000 processed cells.

Extraction and preconcentration of Bisphenol A and 4-Nonylphenol in aqueous solutions using microfunnel supported liquid-phase microextraction prior to high performance liquid chromatography

In this study, liquid-phase microextraction using low-density extraction solvents with microfunnel was applied for the extraction and preconcentration of Bisphenol A and 4-Nonylphenol in aqueous samples. The goal of the present method is to develop a special device that allows organic solvent remain on the surface of aqueous phase as a thin layer during the extraction time. At the end of extraction period, organic phase containing the extracted analytes was collected easily and analyzed by HPLC–FLD. Toluene used as extraction solvent and some of experimental parameters were optimized by L16 Taguchi experimental design. According to the results, the volume of extraction solvent as well as ionic strength showed significant effect on the extraction recovery. Under the optimum conditions (sample volume: 320 mL; pH 8.0; ionic strength: 10% (w/v) NaCl, extraction time: 90 min and extractant: 600 μL toluene), limit of detection, limit of quantification and dynamic linear range of the proposed method for Bisphenol A were calculated as 0.05, 0.2 and 0.2–62.5 μg L−1, and for Nonylphenol were obtained as 3.1, 6.2 and 6.2–125 μg L−1, respectively.

Fuel strand-powered self-propelled electrochemical DNA machine for enzyme-free and distinctly amplified detection of nucleic acid with tunable performance

In this work, a self-propelled DNA machine was proposed for enzyme-free and ultrasensitive electrochemical detection of nucleic acid, which was based on a new strategy of the fuel strand-powered target recycling and successive proximity-based inter-strand displacement for distinct signal amplification. A three-strand duplex DNA (TSD) probe was immobilized onto electrode for target DNA recognition via a fuel strand-powered strand displacement circuit, accompanied with the target recycling and the association of fuel strand onto electrode for signal amplification and readout. The associated fuel strand contained a protruding tail sequence that could be used as a target analogy to activate the neighboring TSD probe based on the proximity-based inter-strand displacement effect, inducing the self-propelled association of fuel strand onto electrode for further signal amplification. The detection performance (dynamic response range, linear interval and detection limit) of current electrochemical DNA machine could be interestingly tuned by changing the tail sequence length of fuel strand, showing the potential for different analysis requirements. The lowest detection limit of 0.1 fM could be achieved, which was lower about two orders of magnitude than that by the typical fuel strand-powered target recycling strategy. Therefore, the developed sensing system exhibits a new, simple and powerful means for amplified detection of nucleic acid and may hold great potentials in bioanalysis, disease diagnosis and biomedicine.

Measurement Device for Ambient Carbonyl Sulfide by Means of Catalytic Reduction Followed by Wet Scrubbing/Fluorescence Detection.

A portable chemical analysis system for monitoring ambient carbonyl sulfide (COS) was investigated for the first time. COS is paid attention to from the perspectives of photosynthesis tracer, breath diagnosis marker, and new process-use in the manufacture of semiconductors. Recently, the threshold level value of COS was settled at 5 ppm in volume ratio (ppmv) for workplace safety management. In this work, COS was converted to H2S by a small column packed with alumina catalyzer at 65 °C. Then, the H2S produced was collected in a small channel scrubber to react with fluorescein mercuric acetate (FMA), and the resulting fluorescence quenching was monitored using an LED/photodiode-based miniature detector. The miniature channel scrubber was re-examined to determine its robustness and easy fabrication, and conditions of the catalyzer were optimized. When the FMA concentration used was 1 μM, the limit of detection and dynamic range, which were both proportional to the FMA concentration, were 0.07 and 25 ppbv, respectively. Ambient COS in the background level and even contaminated COS in the nitrogen gas cylinder could be detected. If necessary, H2S was removed selectively by reproducible adsorbent columns. COS concentrations of engine exhaust were measured by the proposed method and by cryo-trap-gas chromatography-flame photometric detection, and the results obtained (0.5–5.9 ppbv) by the two methods agreed well (R2 = 0.945, n = 19). COS in ambient air and exhaust gases was successfully measured without any batchwise pretreatment.

Smart microfluidic analogue of Wheatstone-bridge for real-time continuous detection with ultrasensitivity and wide dynamic range

Real-time continuous detection of analytes such as environmental toxins is of great significance for myriad applications, ranging from environmental protection to warfare analysis. Efficient signal conversion and amplification are crucial for achieving real-time continuous detection with ultrasensitivity and wide dynamic range. Inspired by the electrical Wheatstone-bridge, a smart microfluidic analogue of Wheatstone-bridge (SMAW) with stimuli-responsive microgel as a variable flow-resistor is developed for such high-performance detection. Similar as current in Wheatstone-bridge, microflows in the SMAW microchip with well-designed microchannel sizes enables new mechanisms to efficiently convert and amplify minute volume changes of microgel induced by trace analytes, into significant width changes of indicating flow as easily measurable output. The typical use of SMAW microchip is highlighted by real-time continuous detection of Pb2+ using a simple optical microscope, with wide dynamic range (10−7 M ~ 10−14 M) and ultra-low detection limit (10−14 M, the lowest one reported to date). This Pb2+-detection also enables fast response, high selectivity, low sample volume, and easy-to-use protocol. The extraordinary SMAW microchip is general and can be extended to detect myriad analytes due to the wide choices of stimuli-responsive hydrogels, thus opening brand-new areas for developing high-performance microfluidic detectors.

Dynamic Range Expansion by Gas-Phase Ion Fractionation and Enrichment for Imaging Mass Spectrometry.

In the analysis of biological tissue by imaging mass spectrometry (IMS), the limit of detection and dynamic range are of paramount importance in obtaining experimental results that provide insight into underlying biological processes. Many important biomolecules are present in the tissue milieu in low concentrations and in complex mixtures with other compounds of widely ranging abundances, challenging the limits of analytical technologies. In many IMS experiments, the ion signal can be dominated by a few highly abundant ion species. On trap-based instrument platforms that accumulate ions prior to mass analysis, these high abundance ions can diminish the detection and dynamic range of lower abundance ions. Herein, we describe two strategies for combating these challenges during IMS experiments on a hybrid QhFT-ICR MS. In one iteration, the mass resolving capabilities of a quadrupole mass filter are used to selectively enrich ions of interest via a technique previously termed continuous accumulation of selected ions. Second, we have introduced a supplemental dipolar AC waveform to the quadrupole mass filter of a commercial QhFT-ICR mass spectrometer to perform selected ion ejection prior to the ion accumulation region. This setup allows the selective ejection of the most abundant ion species prior to ion accumulation, thereby greatly improving the molecular depth with which IMS can probe tissue samples. The gain in sensitivity of both of these approaches roughly scales with the number of accumulated laser shots up to the charge capacity of the ion accumulation cell. The efficiencies of these two strategies are described here by performing lipid imaging mass spectrometry analyses of a rat brain.

Highly Multiplex Targeted Proteomics Enabled by Real-Time Chromatographic Alignment

Targeted mass spectrometry methods produce high-quality quantitative data in terms of limits of detection and dynamic range, at the cost of a substantial compromise in throughput compared to methods such as data independent and data dependent acquisition. The logistical and experimental issues inherent to maintaining assays of even several hundred targets are significant. Prominent among these issues is the drift in analyte retention time as liquid chromatography (LC) columns wear, forcing targeted scheduling windows to be much larger than LC peak widths. If these problems could be solved, proteomics assays would be capable of targeting thousands of peptides in an hour-long experiment, enabling large cohort studies to be performed without sacrificing sensitivity and specificity. We describe a solution in the form of a new method for real-time chromatographic alignment and demonstrate its application to a 56 min LC-gradient HeLa digest assay with 1489 targets. The method is based on the periodic acquisition of untargeted survey scans in a reference experiment and alignment to those scans during subsequent experiments. We describe how the method enables narrower scheduled retention time windows to be used. The narrower scheduling windows enables more targets to be included in the assay or proportionally more time to be allocated to each target, improving the sensitivity. Finally, we point out how the procedure could be improved and how much additional target multiplexing could be gained in the future.

Use of solid phase extraction for sample clean-up and preconcentration of vitamin B12 in multivitamin tablet before HPLC-UV, UV and atomic absorption spectrophotometry

A solid phase extraction method for sample clean-up and preconcentration of vitamin B12 in multivitamin tablet and its determination by HPLC-UV, UV and atomic absorption spectrophotometry is reported. Solid phase parameters such as maximum loading capacity and breakthrough volume were 400 μg and 1750 mL of 0.001 ppm for this vitamin respectively. A proper mobile phase used for HPLC was water/acetonitrile (75:25, v/v) with pH = 3.5 (adjusted with acetate buffer) at 25°C. Limit of detection and linear dynamic range for HPLC, UV and atomic absorption were 0.1, 0.1-25.0; 0.4, 0.4 -15.0; 2.7, 2.7-300.0 (μg/mL) respectively. Recovery of vitamin B12 was about 96%.

Ethanol Determination in Post-Mortem Samples: Correlation between Blood and Vitreous Humor Concentration.

Ethanol (ethylic alcohol) represents the most commonly used drug worldwide and is often involved in clinical and forensic toxicology. Based on several reports, excessive alcohol consumption is the main contributing factor in traffic accidents, drownings, suicides, and other crimes. For these reasons, it becomes essential to analyze the alcohol concentration during autopsy. Although blood is usually used for alcohol analysis in post-mortem cases, it could suffer alterations, putrefaction, and microbial contaminations. As an alternative to whole blood, vitreous humor has been successfully used in medico-legal studies. In this work, post-mortem specimens were analyzed for ethanol determination. The analysis of blood and vitreous humor were carried-out using gas chromatography-flame ionized detector (GC-FID) with a total run time of 6 min. The method was validated in terms of limit of detection, limit of quantification, dynamic range, sensibility, recovery, precision and trueness. A linear regression analysis indicated a coefficient of determination (R2) of 0.9981. The study confirmed no statistically differences between alcohol concentration in blood and vitreous humor, leading vitreous humor as an excellent matrix that could be used as an alternative to whole blood in toxicological analysis in cases where blood is not available.