Detection Of Multiple Analytes Research Articles

Fabrication of a novel mixed-valent copper-based coordination polymer as a fluorescent sensor for selective and efficient detection of multiple analytes

A novel three-dimensional fluorescent mixed-valent copper-based coordination polymer (CP), namely {[CuICuII4(triazol)6(bpta)2]Cl}n (Cu-CP), was synthesized hydrothermally in the presence of Cu(II) ion, 2,4,6-tri(1H-1,2,4-triazol-1-yl)-1,3,5-triazine (TTT), and 2,2′,4,4′-biphenyltetracarboxylic acid (H4bpta). The structure of the Cu-CP was studied using single-crystal X-ray diffraction (SCXRD) and powder X-ray diffraction (PXRD). Interestingly, the crystallographic data revealed that a part of Cu(II) ions were reduced in situ to Cu(I) ions as well as the nitrogen-containing ligand TTT was converted to triazol, which possessed excellent fluorescence characteristics as a transition metal ion with a d10 electronic configuration, prompting further investigation into the fluorescence sensing properties of Cu-CP. The results confirmed that Cu-CP exhibited excellent fluorescence performance which an outstanding candidate as a fluorescence sensor for Fe3+, Cr2O72−, CrO42−, MnO4− and nitrobenzene (NB), with a low limit of detection. For ethylene glycol (EG) detection, Cu-CP showed high-efficiency fluorescence enhancement across a range of temperatures. The underlying mechanism of fluorescence enhancement was investigated through comprehensive experimental studies.

Zr-MOF-based substrate with inherent internal standards facilitates reliable SERS analysis of sulfonamide antibiotics

A multifunctional structure comprising gold nanoparticles immobilized within a zirconium-based metal-organic framework (AuNPs/Zr-MOF) was systematically designed. With the capabilities of analyte enrichment, Raman signal enhancement and internal standard calibration, the as-prepared AuNPs/Zr-MOF substrates demonstrate the outstanding SERS sensing performance toward the representative sulfonamide antibiotics (SAs), sulfadiazine, sulfamerazine, sulfamethazine and sulfaquinoxaline. For the first time, these four SAs are detected by SERS in real honey samples at levels of 2.8 ∼ 5.2 ng·g−1 within 10 min. The extensive π-bond conjugated system of Zr-MOF significantly enhances the concentration of SAs through π-π interactions. The entrapped SAs are adsorbed perpendicularly onto the surface of Au via coordinated interactions between Au and two oxygen atoms, or between Au and the terminal amino group. To achieve reliable quantitative analysis, the inherent vibration of the Zr-MOF structure was used as an internal standard label. Combined with a simple clean-up protocol, the established SERE sensing method is superior to conventional methods in terms of time, while achieving the impressive sensitivity. This study presents a robust SERS-based approach for the rapid detection and monitoring of multiple analytes.

Unlocking multi-mode sensing potential: Phosphorus-doped graphitic carbon nitride quantum dots for Ag+, ciprofloxacin, and riboflavin analysis in environment and food matrices

The simultaneous detection of multiple analytes through a single fluorescence sensor is highly attractive. In this study, phosphorus-doped graphitic carbon nitride quantum dots (P-CNQDs) were developed, achieving multi-mode sensing through three distinct response mechanisms. The preparation involved using melamine as the carbon and nitrogen source and ammonium dihydrogen phosphate as the phosphorus source. Uniform and narrowly distributed P-CNQDs were successfully synthesized through chemical oxidation and hydrothermal methods, with an average size of 2.4 nm. These unique P-CNQDs exhibited fluorescence quenching through photo-induced electron transfer (PET) in response to Ag+. Additionally, the formation of hydrogen bonds and coordination interactions between P-CNQDs-Ag+ and ciprofloxacin (CIP) led to a pronounced fluorescence response to CIP by the chelation enhanced fluorescence (CHEF) mechanism. Furthermore, leveraging the principle of fluorescence resonance energy transfer (FRET), P-CNQDs-CIP served as a ratio fluorescence sensor for riboflavin (RF), enabling ultra-sensitive detection of RF. The combination of PET, CHEF, and FRET response mechanisms successfully facilitated multi-mode sensing for Ag+, CIP, and RF. The detection ranges were 0.05–100 μM, 0.002–2 μM, and 0.05–60 μM, with corresponding lowest detection limits of 17.1 nM, 1.1 nM, and 29.2 nM, respectively. This versatile sensor has been effectively applied to real samples, including the detection of river water and vitamin B2 tablets.

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.

Synthesis of eco-friendly multifunctional dextran microbeads for multiplexed assays

There has been an increasing demand for simultaneous detection of multiple analytes in one sample. Microbead-based platforms have been developed for multiplexed assays. However, most of the microbeads are made of non-biodegradable synthetic polymers, leading to environmental and human health concerns. In this study, we developed an environmentally friendly dextran microbeads as a new type of multi-analyte assay platform. Biodegradable dextran was utilized as the primary material. Highly uniform magnetic dextran microspheres were successfully synthesized using the Shirasu porous glass (SPG) membrane emulsification technique. To enhance the amount of surface functional groups for ligand conjugation, we coated the dextran microbeads with a layer of dendrimers via a simple electrostatic adsorption process. Subsequently, a unique and efficient click chemistry coupling technique was developed for the fluorescence encoding of the microspheres, enabling multiplexed detection. The dextran microbeads were tested for 3-plex cytokine analysis, and exhibited excellent biocompatibility, stable coding signals, low background noise and high sensitivity.

Evolution of 3d printing technology in fabrication of microfluidic devices and biological applications: a comprehensive review

Lab-on-a-chip or LOC is a term that is used to describe microfluidic devices that integrate multiple analyte detection, which are normally carried out in a laboratory, into one micro-chip unit and may have applications in diverse fields such as electronics, medicine and biomedical domains. Even though microfluidics has advanced greatly during the past decade due to increased needs for portability, reduced sample requirement and multiple analyte detection capabilities biological research has not adopted the technology at the required pace. This may be owing to the time-consuming and expensive process involved in the microfabrication of biochips, the requirement of specialised setup facilities and the extremely high cost associated with microfluidics as compared to conventional technologies. In recent years, three-dimensional (3D) printing has piqued curiosity in the scientific community. It has the potential to create complex, high-resolution structures and that too in a short timeframe depending upon device complexity. This could inspire progressive research in microfluidics, particularly finding applications in biomedical engineering and point-of-care diagnostics. This article gives an overview of how 3D printing aids in the manufacture of microfluidic devices for biological applications, as well as the existing 3D printing methods which are utilised for fabrication and the future perspective in the development of microfluidic devices.

Three species multiplexing of fluorescent dyes and gold nanoclusters recovered with fluorescence lifetime correlation spectroscopy

Biosensor applications often require the simultaneous detection of multiple analytes, with a clear need to go beyond the traditional multiplexing relying on distinct fluorescent dyes across the visible spectrum. Fluorescence lifetime correlation spectroscopy (FLCS) is a powerful approach taking advantage of the fluorescence lifetime information to separate the contributions of different fluorescent species with overlapping emission spectra. However, so far FLCS detection has been demonstrated only on binary mixtures of two fluorescent dyes, limiting its multiplexing capabilities. Here, we report the first quantitative FLCS measurements within a ternary mixture composed of three different fluorescent emitters with near-identical emission spectra. Two organic fluorescent dyes, Alexa Fluor 647 and CF640R, are combined with water-soluble Au18(SG)14 gold nanoclusters. Our experimental data establish that FLCS allows to accurately determine individual concentrations within intricate ternary mixtures. Another major aspect of interest concerns the assessment of the suitability of gold nanoclusters for FLCS multiplexing applications. With their microsecond lifetime and stable emission characteristics, gold nanoclusters add a valuable new aspect to the array of FLCS probes. Extending FLCS multiplexing beyond binary mixtures paves the way for further progress in the simultaneous highly parallel biosensing of multiple species.

Abstract 3706: Single cell and cell-free nucleic acid analyses of liquid biopsy specimens from a blood collection tube using a new droplet digital PCR system

Abstract Introduction: Liquid biopsy-based testing has increasingly been adopted by oncologists for biomarker-driven diagnostics, therapy selection, and patient monitoring. Unlike tissue-based approaches, the target analytes in blood specimens are often rare events that must be discriminated from a background of non-tumor derived material. Droplet digital PCR (ddPCR) is an ideal platform for detection of rare nucleic acid markers in the clinic, because it is highly sensitive and specific, robust, and quantitative. Additionally, the workflow is simple to adopt and execute. The QX600 System is a new ddPCR instrument designed for advanced multiplexing which enables simultaneous detection of up to 6 unique fluorophores in a single well. This capability can decrease the sub-sampling needed for detection of multiple analytes. The objective of our study was to develop multiplexed ddPCR assays for detection of cell-based and cell-free circulating biomarkers from a single tube of whole blood. Methods: Tumor cells were enriched from the buffy coat fraction of whole blood collected in fixative-containing cfDNA BCT based on size and morphology using a microfluidics-based system. Enriched tumor cells were enumerated by immunofluorescence (DAPI+/Pan-CK+/CD45-), and gene expression was measured using the QX600 for a panel of cytokeratins (CK) and PD-L1 in CTC negative, NSCLC donor specimens (n=8). Results were compared to CTC positive, contrived specimens (whole blood spiked with breast tumor cell line MCF7 cells; n=5). For ctDNA analysis of the plasma fraction, a multiplexed ddPCR assay was designed to test for five actionable mutations prevalent in NSCLC, including EGFR del19, L858R, T790M, KRAS G12C, BRAF V600E, and wildtype EGFR. To evaluate performance of the multiplexed assay, concordance evaluation was conducted using cfDNA from 8 mutation-positive NSCLC donors on the QX600 and compared to single assays run on the QX200 system. Results: CK8 and CK19 were differentially expressed in the cells enriched from the CTC negative NSCLC cohort as compared to the contrived positives (p=0.0058, and p=0.0049, respectively). All the NSCLC cfDNA samples were positive for the reference variant, and linear regression analysis of the minor variant frequency for ctDNA biomarkers showed good concordance between QX systems (R2=0.8847). Conclusions: These studies demonstrate proof of concept for single cell and cell-free analysis from a single blood draw into a cfDNA BCT using the QX600 system. Liquid biopsy analysis of CTCs and ctDNA can enable minimally invasive, comprehensive decision making in the diagnostic workup of patients. Future studies will focus on further optimizing the cell enrichment methodology from cfDNA BCT and development of additional multiplexed ddPCR assays for NSCLC-relevant biomarkers on the QX600. Citation Format: Leisa Jackson, Colin Cochran, Claire Gould, Sarah Kreston, Jazmin Bello, Brittany D’Alessio, Janice Riley, Alisa Tubbs, Gary A. Pestano. Single cell and cell-free nucleic acid analyses of liquid biopsy specimens from a blood collection tube using a new droplet digital PCR system [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3706.

A Wash-Free Spheres-on-Sphere Strategy for On-Site and Multiplexed Biosensing.

Respiratory infections and food contaminants pose severe challenges to global health and the economy. A rapid on-site platform for the simultaneous detection of multiple pathogens is crucial for accurate diagnosis, appropriate treatment, and a reduced healthcare burden. Herein, we present a spheres-on-sphere (SOS) platform for multiplexed detection using a portable Coulter counter, which employs millimeter- and micron-sized spheres coupled with antibodies as multitarget probes. The assay allows for quantitative detection of multiple analytes within 20 min by simple mixing, enabling on-site detection. The platform shows high accuracy in identifying three respiratory viruses (SARS-CoV-2, influenza A virus, and parainfluenza virus) from throat swab samples, with LOD of 50.7, 32.4, and 49.1 pg/mL. It also demonstrates excellent performance in quantifying three mycotoxins (aflatoxin B1, deoxynivalenol, and ochratoxin A) from food samples. The SOS platform offers a rapid on-site approach with high sensitivity and specificity for applications in resource-limited settings.

Simultaneous SERS Detection of Multiple Amino Acids Using ZIF-8@AuNPs as Substrate: Classified with 1D Convolutional Neural Network

Simultaneous SERS Detection of Multiple Amino Acids Using ZIF-8@AuNPs as Substrate: Classified with 1D Convolutional Neural Network

NIR spectroscopy-CNN-enabled chemometrics for multianalyte monitoring in microbial fermentation.

As the biopharmaceutical industry looks to implement Industry 4.0, the need for rapid and robust analytical characterization of analytes has become a pressing priority. Spectroscopic tools, like near-infrared (NIR) spectroscopy, are finding increasing use for real-time quantitative analysis. Yet detection of multiple low-concentration analytes in microbial and mammalian cell cultures remains an ongoing challenge, requiring the selection of carefully calibrated, resilient chemometrics for each analyte. The convolutional neural network (CNN) is a puissant tool for processing complex data and making it a potential approach for automatic multivariate spectral processing. This work proposes an inception module-based two-dimensional (2D) CNN approach (I-CNN) for calibrating multiple analytes using NIR spectral data. The I-CNN model, coupled with orthogonal partial least squares (PLS) preprocessing, converts the NIR spectral data into a 2D data matrix, after which the critical features are extracted, leading to model development for multiple analytes. Escherichia coli fermentation broth was taken as a case study, where calibration models were developed for 23 analytes, including 20 amino acids, glucose, lactose, and acetate. The I-CNN model result statistics depicted an average R2 values of prediction 0.90, external validation data set 0.86 and significantly lower root mean square error of prediction values ∼0.52 compared to conventional regression models like PLS. Preprocessing steps were applied to I-CNN models to evaluate any augmentation in prediction performance. Finally, the model reliability was assessed via real-time process monitoring and comparison with offline analytics. The proposed I-CNN method is systematic and novel in extracting distinctive spectral features from a multianalyte bioprocess data set and could be adapted to other complex cell culture systems requiring rapid quantification using spectroscopy.

Multiplex Aptamer-Based Fluorescence Assay Using Magnetism-Encoded Nanoparticles for Simultaneous Detection of Multiple Pathogenic Bacteria.

Multiplex assay has emerged as a robust and versatile method for the simultaneous detection of multiple analytes in a single test. However, challenges in terms of poor accuracy and complexity remained. In this work, we developed a multiplex aptamer-based fluorescence assay using magnetism-encoded nanoparticles for the simultaneous detection of multiple pathogenic bacteria. The encapsulation of different amounts of Fe3O4 nanoparticles in zeolitic imidazolate framework-90 (ZIF-90) leads to the formation of Fe3O4@ZIF-90 (FZ) composites with distinct magnetism strengths. By functionalizing a specific aptamer on the surface of the FZ composites, target bacteria can be specifically and precisely separated from a mixed sample in a sequential manner. This property allows for the simultaneous quantitative analysis of multiple target bacteria by using a single-color fluorescence label, thereby resulting in minimal spectral crosstalk interference and improved accuracy. The successful determination of multiple bacteria in contaminated milk samples demonstrates the applicability of this multiplex assay in complex biological matrices. Compared to conventional multiplex fluorescence assays, this approach offers distinct advantages of simplicity, efficiency, and implementation. We believe that this study can provide valuable insights into the development of the multiplex assay while introducing a new method for the simultaneous detection of multiple bacteria.

Discriminative detection of glucose and urea with a composite polymer nanofiber based matrix

AbstractDetection and quantification of glucose and urea are fundamental in medical analysis, real‐time monitoring, and point of care analysis. However, such analyses generally require sophisticated instrumentation and specialized personnel. Furthermore, discriminative detection of multiple analytes is often necessary for treatment and diagnosis. To address this challenge, this study describes a proof‐of‐concept for discriminative sensitive amperometric detection of both analytes. This was achieved by the fabrication of a polymer‐MWCNTs composite nanofiber matrix upon which glucose oxidase (GOD) and urease were co‐immobilized to allow detection of the corresponding substrates (glucose and urea). The (PAN(−MWCNTs)/Urease‐GOD) biosensor showed good performance in: (i) glucose biosensing with sensitivity of 31±5 μAmM−2cm−2 within an extended linear range (0.1–5.0 mM) and a limit of detection of 3.7 μM, and (ii) urea detection at very low concentration (4–30 and 30–90 μM) with sensitivity up to 350±19 μAmM−2cm−2 and very low LOD (0.1 μM). The analogous mono‐enzyme platform for urea detection (PAN(−MWCNTs)/Urease) showed very similar performance as the bi‐enzyme sensor, suggesting little to no influence of co‐immobilization on Urease activity, thus confirming that the two catalytic processes are independent of each other. The biosensors exhibited high accuracy in the detection of both analytes (at normal and pathological levels) in human serum, thus proving that the biosensor described herein can have practical applications in healthcare and remote patient monitoring.

A novel approach to low-cost, rapid and simultaneous colorimetric detection of multiple analytes using 3D printed microfluidic channels.

This research paper presents an inventive technique to swiftly create microfluidic channels on distinct membrane papers, enabling colorimetric drug detection. Using a modified DIY RepRap 3D printer with a syringe pump, microfluidic channels (µPADs) are crafted on a flexible nylon-based substrate. This allows simultaneous detection of four common drugs with a single reagent. An optimized blend of polydimethylsiloxane (PDMS) dissolved in hexane is used to create hydrophobic channels on various filter papers. The PDMS-hexane mixture infiltrates the paper's pores, forming hydrophobic barriers that confine liquids within the channels. These barriers are cured on the printer's hot plate, controlling channel width and preventing spreading. Capillary action drives fluid along these paths without spreading. This novel approach provides a versatile solution for rapid microfluidic channel creation on membrane papers. The DIY RepRap 3D printer integration offers precise control and faster curing. The PDMS-hexane solution accurately forms hydrophobic barriers, containing liquids within desired channels. The resulting microfluidic system holds potential for portable, cost-effective drug detection and various sensing applications.

Non-invasive detection of multiple analytes in dairy industry using heterogeneous ring resonating sensor

Non-invasive detection of multiple analytes in dairy industry using heterogeneous ring resonating sensor

A Microsphere Immunoassay for the Quantitative Detection of Antigens in Cell Culture Supernatant.

Quantitative immunoassays, such as the traditional enzyme-linked immunosorbent assay (ELISA), are used to determine concentrations of an antigen in a matrix of unknown antigen concentration. Magnetic immunoassays, such as the Luminex xMAP technology, allow for the simultaneous detection of multiple analytes and offer heightened sensitivity, specificity, low sample volume requirements, and high-throughput capabilities. Here, we describe a quantitative immunoassay using the Luminex MAGPIX® System to determine the antigen concentration from liquid samples with unknown concentrations. In detail, we describe a newly developed assay for determining production yields of Drosophila S2-produced Marburg virus (MARV) glycoprotein in insect-cell-culture-derived supernatant. The potential applications of this assay could extend to the quantification of viral antigens in fluids derived from both in vitro and in vivo models infected with live MARV, thereby providing additional applications for virological research.

A new indium-phenanthroline based complex as a fluorescence sensor: Synthesis, characterization and detection of multiple analytes

Chemical contaminants have posed a growing danger to environmental security and human health in recent years, causing considerable alarm. Herein, a new indium-based complex was synthesized by solvothermal method and using 1,10-phenanthroline (phen), namely, [(CH3)2NH2]+·[In(phen)Cl4]− (1). The crystal structure of the complex 1 was determined by single crystal X-ray diffraction (SCXRD). Moreover, elemental analysis (EA), Fourier transform infrared spectrum (FTIR), powder X-ray diffraction (PXRD) analysis and other structural characterization were carried out in detail. Thermal and solvent stabilities were investigated thoroughly. The liquid and solid fluorescence spectra had been also explored to discuss the influence of synthesis conditions on crystal configuration and the relationship between the complex and fluorescence properties. Moreover, complex 1 has good fluorescence properties and can be served as an outstanding candidate for the multi-response sensing of Fe3+, Cu2+, Ni2+, Co2+, Cr2O72− ions and nitrobenzene (NB). Furthermore, the limit of detection (LOD) and fluorescence quenching mechanism were studied in detail.

Sweat Sensor Based on Wearable Janus Textiles for Sweat Collection and Microstructured Optical Fiber for Surface-Enhanced Raman Scattering Analysis.

Wearable sweat sensors provide real-time monitoring of biomarkers, enabling individuals to gain real-time insight into their health status. Current sensors primarily rely on electrochemical mechanisms, limiting their capacity for the concurrent detection of multiple analytes. Surface-enhanced Raman scattering spectroscopy offers an alternative approach by providing molecular fingerprint information to facilitate the identification of intricate analytes. In this study, we combine a wearable Janus fabric for efficient sweat collection and a grapefruit optical fiber embedded with Ag nanoparticles as a sensitive SERS probe. The Janus fabric features a superhydrophobic side in contact with the skin and patterned superhydrophilic regions on the opposite surface, facilitating the unidirectional flow of sweat toward these hydrophilic zones. Grapefruit optical fibers feature sharp tips with the ability to penetrate transparent dressings. Its microchannels extract sweat through capillary force, and nanoliter-scale volumes of sweat are sufficient to completely fill them. The Raman signal of sweat components is greatly enhanced by the plasmonic hot spots and accumulates along the fiber length. We demonstrate sensitive detection of sodium lactate and urea in sweat with a detection limit much lower than the physiological concentration levels. Moreover, the platform shows its capability for multicomponent detection and extends to the analysis of real human sweat.

Gold nanoparticles as colorimetric probes in food analysis: Progress and challenges.

The rapid, sensitive and reliable food safety control is urgently needed due to the harmful effects of the food contaminants on human health. Colorimetric approach has exhibited promising potential for the detection of food contaminants due to their easy preparation, rapid detection, high sensitivity, and naked-eye sensing. In recent years, AuNPs-based colorimetric probes have been extensively explored for food analysis. The present article reviews the development of AuNPs-based colorimetric probes for colorimetric sensing and their applications in food analysis. It generally summarizes the properties of AuNPs and introduces the preparation and functionalization methods of AuNPs. An overview of the colorimetric sensing mechanisms of AuNPs-based probes and their applications in analysis of food contaminants are also provided. Although AuNPs-based colorimetric probes show many advantages in detection of food contaminants, challenges remain in terms of complexity of food matrices, multiple analytes detection in a single go, and testing conditions interference.

Design and analysis of a high sensitivity open microchannel PCF-based surface plasmon resonance refractometric sensor

This paper presents a high-sensitivity dual-open microchannel PCF-SPR refractometric sensor that was numerically analyzed using the Finite Element Method (FEM). A dual-open microchannel was created on both sides of the fiber to facilitate analyte infiltration and improve its detection performance. The microchannel of PCF was coated with gold plasmonic material to achieve practical sensing. The results revealed that the refractive index sensing range of the proposed refractometric sensor is between 1.37 and 1.43, with a highest amplitude sensitivity of 3743.64 RIU−1 and a wavelength sensitivity of 33000 nm/RIU. Additionally, it offers a maximum resolution of 5.56 × 10−6 RIU. All structure parameters, such as air-hole length, pitch, open-channel radius, and gold thickness, are optimized. The proposed sensor is a significant improvement in the detection of multiple analytes in bio-sensing applications.