A Separation‐Sensing Platform Performing Accurate Diagnosis of Jaundice in Complex Biological Tear Fluids

The detection of biomarkers in tears has aroused great interest owing to the advantages of non‐invasive and rapid collection. The combination of ultrasensitivity and label‐free detection of surface‐enhanced Raman spectroscopy (SERS) sensors is expected to achieve real‐time diagnosis in home medical care. However, the surface of SERS sensors is susceptible to biofouling and inactivation by biological impurities in tears, resulting in rapid degradation of sensitivity, limiting the commercialization of point‐of‐care devices. Herein, a binary nanosphere array with dual properties is constructed as a separation‐sensing platform for the diagnosis of target molecules in tears. The upper part of the structure is composed of Au nanoparticles (AuNPs) and a sputtering Au layer, which can bind the target molecules that interact with Au and provide high‐strength and high‐density SERS hotspots. The lower half is an inactive SiO2 nanosphere array with periodic large pores that allows biological impurities to penetrate the lower part and be separated from the target analyte. Furthermore, this substrate was integrated into homemade tear kits, enabling simultaneous tear collection, pre‐separation, and detection. Combined with the Raman spectra of tears and LDA analysis, we successfully identified patients with jaundice in clinics. This platform is expected to provide an opportunity for early disease screening based on biological fluids.

The precise, simultaneous, and rapid detection of essential biomarkers in human tears is imperative for monitoring both ocular and systemic health. The utilization of a wearable colorimetric biochemical sensor exhibits potential in achieving swift and concurrent detection of pivotal biomarkers in tears. Nevertheless, challenges arise in the collection, interpretation, and sharing of data from the colorimetric sensor, thereby restricting the practical implementation of this technology. To overcome these challenges, this research introduces an artificial intelligence-assisted wearable microfluidic colorimetric sensor system (AI-WMCS) for rapid, non-invasive, and simultaneous detection of key biomarkers in human tears, including vitamin C, H+(pH), Ca2+, and proteins. The sensor consists of a flexible microfluidic epidermal patch that collects tears and facilitates the colorimetric reaction, and a deep-learning neural network-based cloud server data analysis system (CSDAS) embedded in a smartphone enabling color data acquisition, interpretation, auto-correction, and display. To enhance accuracy, a well-trained multichannel convolutional recurrent neural network (CNN-GRU) corrects errors in the interpreted concentration data caused by varying pH and color temperature in different measurements. The test set determination coefficients (R2) of 1D-CNN-GRU for predicting pH and 3D-CNN-GRU for predicting the other three biomarkers were as high as 0.998 and 0.994, respectively. This correction significantly improves the accuracy of the predicted concentration, enabling accurate, simultaneous, and quick detection of four critical tear biomarkers using only minute amounts of tears ( ~ 20 μL). This research demonstrates the powerful integration of a flexible microfluidic colorimetric biosensor and deep-learning algorithm, which holds immense potential to revolutionize the fields of health monitoring.

Automation Highlights from the Literature

BackgroundThere is a long-lasting need for non-invasive, more accurate diagnostic techniques when evaluating primary Sjögren’s syndrome (pSS) patients. Incorporation of additional diagnostics involving screening for disease-specific biomarkers in biological fluid is a promising concept that requires further investigation. In the current study we aimed to explore novel disease biomarkers in saliva and tears from pSS patients.MethodsLiquid chromatography-mass spectrometry (LC-MS) was performed on stimulated whole saliva and tears from 27 pSS patients and 32 healthy controls, and salivary and tear proteomic biomarker profiles were generated. LC-MS was also combined with size exclusion chromatography to isolate extracellular vesicles (EVs) from both fluids. Nanoparticle tracking analysis was conducted on joint fractions from the saliva and tears to determine size distribution and concentration of EVs. Further EV characterisation was performed by immunoaffinity capture of CD9-positive EVs using magnetic beads, detected by flow cytometry. The LC-MS data were analysed for quantitative differences between patient and control groups using Scaffold, and the proteins were further analysed using the Database for Annotation, Visualization and Integrated Discovery (DAVID), for gene ontology overrepresentation, and the Search Tool for the Retrieval of Interacting Genes/Proteins for protein-protein interaction network analysis.ResultsUpregulation of proteins involved in innate immunity (LCN2), cell signalling (CALM) and wound repair (GRN and CALML5) were detected in saliva in pSS. Saliva EVs also displayed biomarkers critical for activation of the innate immune system (SIRPA and LSP1) and adipocyte differentiation (APMAP). Tear analysis indicated overexpression of proteins involved in TNF-α signalling (CPNE1) and B cell survival (PRDX3). Moreover, neutrophil gelatinase-associated lipocalin was upregulated in saliva and tears in pSS. Consistently, DAVID analysis demonstrated pathways of the adaptive immune response in saliva, of cellular component assembly for saliva EVs, and of metabolism and protein folding in tears in pSS patients.ConclusionsLC-MS of saliva and tears from pSS patients, solely and in combination with size-exclusion chromatography allowed screening for possible novel biomarkers encompassing both salivary and lacrimal disease target organs. This approach could provide additional diagnostic accuracy in pSS, and could possibly also be applied for staging and monitoring the disease.

Biosensor for electrochemical detection of biomarkers of osteoarthritis

1. Introduction -Definitions -Historical aspects of biomarkers -Classification of biomarkers -Types of biomarkers -The ideal biomarker -Biomarkers and systems biology -Relation of biomarkers to other technologies and healthcare 2. Technologies for Discovery of Biomarkers -Introduction -Detection of biomarkers in tissues and body fluids -Disease biomarkers in breath -Genomic technologies -Epigenomic technologies -Proteomic technologies -Glycomic technologies -Metabolomic technologies -Lipidomics -Fluorescent indicators for biomarkers -Molecular imaging technologies -Nuclear magnetic resonance -Nanobiotechnology -Bioinformatics -Pitfalls in the discovery and development of biomarkers 3. Biomarkers and Molecular Diagnostics -Introduction -Molecular diagnostic technologies -Detection and expression profiling of miRNA 4. Biomarkers for Drug Discovery & Development -Introduction -Biomarker technologies for drug discovery -Biomarkers and drug safety -Applications of biomarkers for drug development 5. Role of Biomarkers in Healthcare -Introduction -Biomarkers of inflammation -Biomarkers of oxidative stress -Biomarkers in metabolic disorders -Biomarkers in immune disorders -Biomarkers of musculoskeletal disorders -Biomarkers of osteoporosis -Biomarkers of infectious diseases -Biomarkers of liver disease -Biomarkers of pancreatitis -Biomarkers of renal disease -Biomarkers of pulmonary diseases -Biomarkers in obstetrics and gynecology -Biomarkers for genetic disorders -Biomarkers of aging -Biomarkers of miscellaneous disorders -Biomarkers and nutrition -Biomarkers of gene-environmental interactions in human disease -Future role of biomarkers in healthcare -Applications of biomarkers beyond healthcare 6. Biomarkers of Cancer -Introduction -Types of cancer biomarkers -Molecular diagnostic techniques for cancer -Technologies for detection of cancer biomarkers -Applications of cancer biomarkers -Role of biomarkers in drug development in oncology -Biomarkers according to location/type of cancer -Role of the NCI in biomarkers of cancer -Future prospects for cancer biomarkers 7. Biomarkers of Disorders of the Nervous System -Introduction -Discovery of biomarkers for neurological disorders -Data mining for biomarkers of neurological disorders -Antibodies as biomarkers in disorders of the nervous system -Biomarkers of neural regeneration -Biomarkers of disruption of blood-brain barrier -Biomarkers of neurotoxicity -Biomarkers of neurodegenerative disorders -Biomarkers of multiple sclerosis -Biomarkers of stroke -Biomarkers of traumatic brain injury -Biomarkers of CNS infections -Biomarkers of epilepsy -Biomarkers of normal pressure hydrocephalus -Biomarkers of retinal disorders -Biomarkers for autism -Biomarkers of sleep disorders -Biomarkers of psychiatric disorders 8. Biomarkers of Cardiovascular Disorders -Introduction -Biomarkers of cardiovascular diseases -Methods for identification of cardiovascular biomarkers -Applications of biomarkers of cardiovascular disease -Role of biomarkers in the management of cardiovascular disease -Fut

The detection and monitoring of biomarkers in body fluids has been used to improve human healthcare activities for decades. In recent years, researchers have focused their attention on applying the point-of-care (POC) strategies into biomarker detection. The evolution of mobile technologies has allowed researchers to develop numerous portable medical devices that aim to deliver comparable results to clinical measurements. Among these, optical-based detection methods have been considered as one of the common and efficient ways to detect and monitor the presence of biomarkers in bodily fluids, and emerging aggregation-induced emission luminogens (AIEgens) with their distinct features are merging with portable medical devices. In this review, the detection methodologies that use optical measurements in the POC systems for the detection and monitoring of biomarkers in bodily fluids are compared, including colorimetry, fluorescence and chemiluminescence measurements. The current portable technologies, with or without the use of smartphones in device development, that are combined with optical biosensors for the detection and monitoring of biomarkers in body fluids, are also investigated. The review also discusses novel AIEgens used in the portable systems for the detection and monitoring of biomarkers in body fluid. Finally, the potential of future developments and the use of optical detection-based portable devices in healthcare activities are explored.

Rotational actuation of magnetic nanoparticle clusters for solution-based biosensing

Cancer and neurological disorders are two leading causes of human death. Their early diagnoses will either greatly improve the survival rate or facilitate effective treatments or modalities. Detection of biomarkers in body fluids and some tissues (e.g., blood, urine and cerebrospinal fluids) is relatively non-invasive and provides useful chemical and biological information that is complementary to tomographic imaging (e.g., magnetic resonance imaging, positron emission tomography and X-ray computed tomography). Recent years have witnessed the contributions from and potential applications of bioanalytical methods for early detection of major diseases. In this review, we survey some recent developments of electroanalytical (as a representative label-based technique) and surface plasmon resonance (SPR) (as a representative label-free technique) biosensors for detection of biomarkers relevant to etiologies of breast cancer and Alzheimer's disease (AD). While breast cancer is representative of cancers of complexity (multiple biomarkers, false positives from tomographic scans, and a need for more effective early diagnostic methods), AD is the most prevalent neurological disorder that is also linked to multiple biomarkers. Both electroanalytical and SPR-based sensors have attractive features of sensitivity, portability, obviation of large sample volumes, and capability of multiplexed detection. Various sensing protocols developed in the past five years are reviewed, demonstrating the feasibility of both techniques for diagnostic purposes. Problems inherent in these two techniques that must be overcome before being clinically viable are also discussed.

Extracellular vesicles (EVs) are cell-secreted, lipid membrane-enclosed nanoparticles without functional nucleus. EV is a general term that includes various subtypes of particles named microvesicles, microparticles, ectosomes or exosomes. EVs transfer RNA, DNA and protein cargo between proximal and distant cells and tissues, thus constituting an organism-wide signal transduction network. Pathological tissues secrete EVs that differ in their cargo composition compared to their healthy counterparts. The detection of biomarkers in EVs from biological fluids may aid the diagnosis of disease and/or monitor its progression in a minimally invasive manner. Among biological fluids, pleural effusions (PEs) are integrated to clinical practice, as they accompany a wide variety of lung disorders. Due to the proximity with the pleura and the lungs, PEs are expected to be especially enriched in EVs that originate from diseased tissues. However, PEs are among the least studied biofluids regarding EV-specialized isolation methods and related biomarkers. Herein, we describe a practical EV isolation method from PEs for the screening of EV RNA biomarkers in clinical routine. It is based on a Proteinase K treatment step to digest contaminants prior to standard polyethylene-glycol precipitation. The efficiency of the method was confirmed by transmission electron microscopy, nanoparticle tracking analysis and Western blot. The reliability and sensitivity of the method towards the detection of EV-enriched RNA biomarkers from multiple PEs was also demonstrated.

Biosensor approaches on the diagnosis of neurodegenerative diseases: Sensing the past to the future

The aim of the work: to optimize the method for Trazodone isolation from blood and urine by liquid-liquid extraction technique. Materials and Methods. The study was carried out with model samples of human biofluids spiked with Trazodone. The antidepressant was isolated from the blood and urine by the liquid-liquid extraction technique with methylene chloride from the alkaline aqueous medium at pH 9 at the presence of ammonium sulphate as a salting-out agent. The biological impurities were previously removed by extraction with diethyl ether from the acidic medium at pH 1. The erythrocyte mass of the blood was pre-precipitated with the help of 5% trichloroacetic acid. The resulting extracts were additionally purified using TLC technique. Quantitative determination of Trazodone in the extracts was carried out with the help of UV spectrophotometry. Results and Discussion. Trazodone was detected in the biological extracts by UV spectra which coincided with standard methanol solution of the analyte and had the principal peak at wavelength of 251±2 nm. Quantitative determination of the drug in the extracts was performed with the help of calibration curve described by the equation y=(0.0230±1×10-4)×x, which showed linearity in the range of analyte concentration 2.5–50.0 μg/mL. Recovery values of the developed sample preparation methods were determined in the range of the drug concentrations of 10–50 µg/mL and 4–20 µg/mL for blood and urine respectively. Conclusions. Effective methods for Trazodone isolation from blood and urine using liquid-liquid extraction in the presence of ammonium sulphate as a salting-out agent have been developed. It has been established that liquid-liquid extraction in the presence of electrolytes provided more effective Trazodone isolation from the biological fluids than under optimized conditions without salting-out agents. The recovery of the optimized methods was 52±3% for blood and 91±3% for urine that provided sufficient extraction yield of the antidepressant for its determination within the level of the expected acute concentrations in the biological fluids. The ability of UV spectrophotometry as an analytical method for determination of Trazodone in biological fluids has been proven by a range of validation parameters. The obtained results can be used for forensic toxicological examinations in cases of acute and fatal intoxications by antidepressants.

Ultra-low concentration detection of DNA without its amplification using a biological nanopore.

Identification and detection of protein markers to differentiate between forensically relevant body fluids

In the field of predictive, preventive and personalised medicine, researchers are keen to identify novel and reliable ways to predict and diagnose disease, as well as to monitor patient response to therapeutic agents. In the last decade alone, the sensitivity of profiling technologies has undergone huge improvements in detection sensitivity, thus allowing quantification of minute samples, for example body fluids that were previously difficult to assay. As a consequence, there has been a huge increase in tear fluid investigation, predominantly in the field of ocular surface disease. As tears are a more accessible and less complex body fluid (than serum or plasma) and sampling is much less invasive, research is starting to focus on how disease processes affect the proteomic, lipidomic and metabolomic composition of the tear film. By determining compositional changes to tear profiles, crucial pathways in disease progression may be identified, allowing for more predictive and personalised therapy of the individual.This article will provide an overview of the various putative tear fluid biomarkers that have been identified to date, ranging from ocular surface disease and retinopathies to cancer and multiple sclerosis. Putative tear fluid biomarkers of ocular disorders, as well as the more recent field of systemic disease biomarkers, will be shown.