On-Site Bioaerosol Sampling and Airborne Microorganism Detection Technologies.

Bioaerosols are small airborne particles composed of microbiological fragments, including bacteria, viruses, fungi, pollens, and/or by-products of cells, which may be viable or non-viable wherever applicable. Exposure to these agents can cause a variety of health issues, such as allergic and infectious diseases, neurological disorders, and cancer. Therefore, detecting and identifying bioaerosols is crucial, and bioaerosol sampling is a key step in any bioaerosol investigation. This review provides an overview of the current bioaerosol sampling methods, both passive and active, as well as their applications and limitations for rapid on-site monitoring. The challenges and trends for detecting airborne microorganisms using molecular and immunological methods are also discussed, along with a summary and outlook for the development of prompt monitoring technologies.

AuNPs- Aß-Ni-HRP sandwich assay: A new sensitive colorimetric method for the detection of Aß 1-40.

Amyloid ß-peptide (Aß) is a key predictor for preclinical diagnosis of Alzheimer's disease (AD) and vascular diseases. In this work, we propose a gold nanoparticle (AuNPs)-Aß-nickel (Ni)-horseradish peroxidase (HRP) based colorimetric assay for the detection of Aß1-40. The consecutive binding of Aß1-40 to AuNPs and metal ions is designed and examined for Aß-specific aggregation of AuNPs and the generation of quantitative colorimetric signals. The affinity of Aß1-40 towards various metal ions was studied first, and two metal ions, Cu and Ni, were specifically tested with Metal Ion-Binding Site Prediction (MIB) and High-resolution Electrospray Ionization Mass Spectrometry (HR-ESI-MS). Subsequently, the binding of Aß1-40 and AuNPs was examined, and the binding between Aß-AuNPs and Ni-HRP was finally analyzed by UV-Vis and nano-zetasizer. Based on the characterized dual binding of Aß1-40, a colorimetric sandwich assay was developed and the analytical performance of the developed assay has been evaluated with standard solutions and human serum samples. Good linearity within a range from 0 nM to 10 nM was found. The detection limits of 0.22 nM in the standard sample and 0.23 nM in the human serum sample have been demonstrated. The newly developed colorimetric sandwich assay is a short, simple, antibody-free assay and achieves high sensitivity with only 100 µL Aß1-40 samples. The assay has immense potential for the detection of Aß1-40 in biological or biomedical diagnosis.

An Electrochemical Chip to Monitor In Vitro Glycation of Proteins and Screening of Antiglycation Potential of Drugs.

Hyperglycemia and the production of advanced glycation end products (AGEs) are the primary factors for the development of chronic complications in diabetes. The level of protein glycation is proportional to the glucose concentration and represents mean glycemia. In this study, we present an electrochemical chip-based method for in vitro glycation monitoring and the efficacy evaluation of an antiglycation compound. An electrochemical chip consisting of five microchambers and embedded microelectrodes was designed for parallel measurements of capacitance signals from multiple solutions at different concentrations. The feasibility of glycation monitoring was then investigated by measuring the capacitance signal at 0.13 MHz with bovine serum albumin and gelatin samples in the presence of various glucose concentrations over 28 days. A significant change in the capacitance due to protein glycation was observed through measurements conducted within 30 s and 21 days of incubation. Finally, we demonstrated that the chip-based capacitance measurement can be utilized for the selection of an antiglycation compound by supplementing the protein solution and hyperglycemic concentration of glucose with an inhibitory concentration of the standard antiglycation agent aspirin. The lack of a significant change in the capacitance over 28 days proved that aspirin is capable of inhibiting protein glycation. Thus, a strong relationship exists between glycation and capacitance, suggesting the application of an electrochemical chip for evaluating glycation and novel antiglycation agents.

Detection of C-Reactive Protein Using Histag-HRP Functionalized Nanoconjugate with Signal Amplified Immunoassay.

Ultrasensitive detection of biomarkers is highly significant for disease prognosis and public health treatment. Despite wide acceptance in routine laboratory tests, the conventional enzyme-linked immunosorbent assay (ELISA) has been of limited use for early biomarker detection due to insufficient sensitivity and multiple long incubation time. Several nanoprobes have been introduced to circumvent the limitation, however, rapid, simple, and chemical-free nanoprobe synthesis and sensitive detection methods, particularly for ELISA, are still lacking. In this study, we have synthesized a gold nanoprobe, conjugated with multiple 6X-histidine (6X-his) peptide and nickel-horseradish peroxidase (Ni2+-HRP), for enhancing the colorimetric signal in ELISA. The developed nanoprobe has been tested for the detection of immunologically significant C-reactive protein (CRP) in ELISA format. The performance of designed probe is validated by testing standard and serum samples, and the detection limit of 32.0 pg/mL with R2 = 0.98 is confirmed. Furthermore, a comparative analysis of the developed nanoprobe was performed with ELISA developed on conventional guidelines, the proposed immunoassay showed an increase of 12-fold sensitivity for detecting CRP due to the high loading of 6Xhis peptide and binding of multiple Ni2+-HRP on a gold nanoparticle. Additionally, the proposed assay provides a simple, fast, and cost-efficient (not requiring multiple antibodies) detection of CRP with easy nanoprobe synthesis. Moreover, the developed Histag-HRP functionalized nanoconjugate immunoassay is flexible and can be applied to other biomarkers efficiently by using disease specific antibody.

Label-Free Electrochemical Microfluidic Chip for the Antimicrobial Susceptibility Testing.

The emergence and spread of antibiotic-resistant bacteria is a global threat to human health. An accurate antibiotic susceptibility test (AST) before initiating the treatment is paramount in the treatment and bacterial resistance control. However, the current AST methods either are complex, use chemical and biological labels, lack multiplexing, are expensive, or are too slow to be used for routine screening. The primary objective of the current study is to develop an automated electrochemical microfluidic chip (EMC) for simple and rapid AST. The microfluidic channels and gold microelectrodes were designed for the automation of antibiotic mixing and distribution in multiple test chambers and for electrical signal measurements. The designed chip was tested for AST with E. coli samples, and the results were compared with conventional broth microdilution. The presented EMC provided rapid bacterial count and AST in 170 and 150 min, respectively, while the conventional broth microdilution evaluates in 450 and 240 min, respectively. The rapid AST capability of the EMC was further demonstrated with the artificial urine samples, and the results were obtained in 270 min, which was 90 min faster than the broth microdilution method. Additionally, the minimum inhibitory concentration (MIC) was evaluated on the EMC and compared with the results from an AlamarBlue assay. The experimental results indicate the sensitivity of the chip, minimum loss of antibiotics, and eventually, reduction in the evolution of antibiotic resistance. Cumulatively, we have developed an automated, label-free, economical, rapid, robust, and user-friendly EMC for the evaluation of AST in urine samples.

SPE based soil processing and aptasensor integrated detection system for rapid on site screening of arsenic contamination in soil.

Rapid industrialization and urbanization have resulted in serious environmental deterioration, especially in terms of heavy metal contamination in soil. Arsenic is one of the primary heavy metal contaminants in the soil and possesses a severe threat to all the plants and animals including humans. The conventional methods for analyzing arsenic contamination in soil have tedious, time-consuming sample preparation steps and require laboratory equipped instruments and skilled personnel. The present work demonstrates a novel method for arsenic As(III) detection in the contaminated soil based on field applicable sample preparation and smartphone-based optical sensing. Soil sample preparation has been simplified and optimized using acid extraction and serial application of different solid phase extraction (SPE) cartridges for the removal of interfering ions with high arsenic yield in one step. The acidic extraction and SPE efficiencies were found to be 35.4% and 54.0%, respectively, for arsenic contaminated field soil samples. The quantification of As(III) was performed by aptamer-AuNPs based colorimetric assay with a smartphone coupled optical unit. This aptasensor integrated detection system (ADS) has shown a detection limit of 14.44 ppb for aqueous samples and 1.97 ppm for field soil samples. In the accuracy comparison with ICP-MS, arsenic contaminated field soils from various sources have been tested and the results depicted a highly significant correlation coefficient of 0.997 with an average difference of 1.67 ppm. By integrating all the required analytical steps into a portable format, the presented setup enables on-site tests of arsenic contamination in soil.

Recent advances in high-sensitivity detection methods for paper-based lateral-flow assay.

Paper-based lateral-flow assays (LFAs) have achieved considerable commercial success and continue to have a significant impact on medical diagnostics and environmental monitoring. Conventional LFAs are typically performed by examining the color changes in the test bands by naked eye. However, for critical biochemical markers that are present in extremely small amounts in the clinical specimens, this readout method is not quantitative, and does not provide sufficient sensitivity or suitable detection limit for a reliable assay. Diverse technologies for high-sensitivity LFA detection have been developed and commercialization efforts are underway. In this review, we aim to provide a critical and objective overview of the recent progress in high-sensitivity LFA detection technologies, which involve the exploitation of the physical and chemical responses of transducing particles. The features and biomedical applications of the technologies, along with future prospects and challenges, are also discussed.

Tannic acid-mediated rapid layer-by-layer deposited non-leaching silver nanoparticles hybridized cellulose membranes for point-of-use water disinfection.

Gravity driven water filtration is a commonly used process of removing microorganism from the contaminated water. However, the existing strategies involve prolonged synthesis and toxic reducing agent for immobilization of silver nanoparticles (AgNPs) on cellulose membranes that are not suitable for routine handling. We have developed a non-toxic and environmentally benign method using TA mediated silver salt layer-by-layer (LbL) in-situ reduction method. Our LbL method exhibited the properties of controlled size and uniform distribution of in-site AgNPs on the surface of the membranes. The LbL deposited AgNPs hybrid membranes displayed an excellent antibacterial activity which have been validated through an efficient bacterial filtration performance against the Escherichia coli (E. coli). The present method for developing hybrid membranes offered a simple, rapid, low-cost, sustainable, and large-scale fabrication for bacterial filtration which could be used for the point-of-use applications, particularly at resource-limited and remote areas.

Current and Emerging Methods of Antibiotic Susceptibility Testing.

Antibiotic susceptibility testing (AST) specifies effective antibiotic dosage and formulates a profile of empirical therapy for the proper management of an individual patient's health against deadly infections. Therefore, rapid diagnostic plays a pivotal role in the treatment of bacterial infection. In this article, the authors review the socio-economic burden and emergence of antibiotic resistance. An overview of the phenotypic, genotypic, and emerging techniques for AST has been provided and discussed, highlighting the advantages and limitations of each. The historical perspective on conventional methods that have paved the way for modern AST like disk diffusion, Epsilometer test (Etest), and microdilution, is presented. Several emerging methods, such as microfluidic-based optical and electrochemical AST have been critically evaluated. Finally, the challenges related with AST and its outlook in the future are presented.

Progress in antibiotic susceptibility tests: a comparative review with special emphasis on microfluidic methods.

Antibiotic susceptibility test (AST) is an umbrella term for techniques to determine the susceptibility of bacteria to antibiotics. The antibiotic-resistant bacteria are a major threat to public health and a directed therapy based on accurate AST results is paramount in resistance control. Here we have briefly covered the progress of conventional, molecular, and automated AST tools and their limitations. Various aspects of microfluidic AST such as optical, electrochemical, colorimetric, and mechanical methods have been critically reviewed. We also address the future requirements of the microfluidic devices for AST. Cumulatively, we have outlined the overview of AST that can help to expand and improve the existing techniques and emphasize that microfluidics could be the future of AST and introduction of microtechnologies in AST will be extremely advantageous, especially for point-of-care testing.

Miniaturized Sample Preparation and Rapid Detection of Arsenite in Contaminated Soil Using a Smartphone.

Conventional methods for analyzing heavy metal contamination in soil and water generally require laboratory equipped instruments, complex procedures, skilled personnel and a significant amount of time. With the advancement in computing and multitasking performances, smartphone-based sensors potentially allow the transition of the laboratory-based analytical processes to field applicable, simple methods. In the present work, we demonstrate the novel miniaturized setup for simultaneous sample preparation and smartphone-based optical sensing of arsenic As(III) in the contaminated soil. Colorimetric detection protocol utilizing aptamers, gold nanoparticles and NaCl have been optimized and tested on the PDMS-chip to obtain the high sensitivity with the limit of detection of 0.71 ppm (in the sample) and a correlation coefficient of 0.98. The performance of the device is further demonstrated through the comparative analysis of arsenic-spiked soil samples with standard laboratory method, and a good agreement with a correlation coefficient of 0.9917 and the average difference of 0.37 ppm, are experimentally achieved. With the android application on the device to run the experiment, the whole process from sample preparation to detection is completed within 3 hours without the necessity of skilled personnel. The approximate cost of setup is estimated around 1 USD, weight 55 g. Therefore, the presented method offers the simple, rapid, portable and cost-effective means for onsite sensing of arsenic in soil. Combined with the geometric information inside the smartphones, the system will allow the monitoring of the contamination status of soils in a nation-wide manner.

Advances in microfluidic PCR for point-of-care infectious disease diagnostics.

Global burdens from existing or emerging infectious diseases emphasize the need for point-of-care (POC) diagnostics to enhance timely recognition and intervention. Molecular approaches based on PCR methods have made significant inroads by improving detection time and accuracy but are still largely hampered by resource-intensive processing in centralized laboratories, thereby precluding their routine bedside- or field-use. Microfluidic technologies have enabled miniaturization of PCR processes onto a chip device with potential benefits including speed, cost, portability, throughput, and automation. In this review, we provide an overview of recent advances in microfluidic PCR technologies and discuss practical issues and perspectives related to implementing them into infectious disease diagnostics.

Continuous dielectrophoretic bacterial separation and concentration from physiological media of high conductivity.

Biological sample processing involves purifying target analytes from various sample matrices and concentrating them to a small volume from a large volume of crude sample. This complex process is the major obstacle for developing a microfluidic diagnostic platform. In this study, we present a microfluidic device that can continuously separate and concentrate pathogenic bacterial cells from complex sample matrices such as cerebrospinal fluid and whole blood. Having overcome critical limitations of dielectrophoretic (DEP) operation in physiological media of high conductivity, we utilized target specific DEP techniques to incorporate cell separation, medium exchange, and target concentration into an integrated platform. The proposed microfluidic device can uptake mL volumes of crude biological sample and selectively concentrate target cells into a submicrolitre volume, providing ~10(4) fold of concentration. We designed the device based on the electrokinetic theory and electric field simulation, and tested the device performance with different sample types. The separation efficiency of the device was as high as 97.0% for a bead mixture in TAE buffer and 94.3% and 87.2% for E. coli in human cerebrospinal fluid and blood, respectively. A capture efficiency of 100% was achieved in the concentration chamber. With a relatively simple configuration, the proposed device provides a robust method of continuous sample processing, which can be readily integrated into a fully automated microfluidic diagnostic platform for pathogen detection and quantification.

A surface topography assisted droplet manipulation platform for biomarker detection and pathogen identification.

This paper reports a droplet microfluidic, sample-to-answer platform for the detection of disease biomarkers and infectious pathogens using crude biosamples. The platform exploited the dual functionality of silica superparamagnetic particles (SSP) for solid phase extraction of DNA and magnetic actuation. This enabled the integration of sample preparation and genetic analysis within discrete droplets, including the steps of cell lysis, DNA binding, washing, elution, amplification and detection. The microfluidic device was self contained, with all reagents stored in droplets, thereby eliminating the need for fluidic coupling to external reagent reservoirs. The device incorporated unique surface topographic features to assist droplet manipulation. Pairs of micro-elevations were created to form slits that facilitated efficient splitting of SSP from droplets. In addition, a compact sample handling stage, which integrated the magnet manipulator, the droplet microfluidic device and a Peltier thermal cycler, was built for convenient droplet manipulation and real-time detection. The feasibility of the platform was demonstrated by analysing ovarian cancer biomarker Rsf-1 and detecting Escherichia coli with real time polymerase chain reaction and real time helicase dependent amplification.

Negative dielectrophoretic capture of bacterial spores in food matrices.

A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S∕m), apple juice (0.225 S∕m), and milk (0.525 S∕m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 µm height DEP chambers, "almost complete" and rapid particle capture from lower conductivity (0.05 S∕m) medium is observed. Using DEP chambers above 150 µm in height, the onset of a global fluid motion for high conductivity media is observed. This motion enhances particle capture on the electrodes at the center of the DEP chamber. The n-DEP electrodes are designed to have well defined electric field minima, enabling sample concentration at 1000 distinct locations within the chip. The electrode design also facilitates integration of immunoassay and other surface sensors onto the particle capture sites for rapid detection of target micro-organisms in the future.

An all-in-one microfluidic device for parallel DNA extraction and gene analysis.

We have developed a microfluidic device capable of fully integrated sample preparation and gene analysis from crude biosamples such as whole blood. Our platform takes the advantage of the silica superparamagnetic particle based solid phase extraction to develop an all-in-one scheme that performs cell lysis, DNA binding, washing, elution and the PCR in the same reaction chamber. The device also employs a unique reagent loading scheme, allowing efficient preparation of multiple reactions via a single injection channel. In addition, PCR is performed in a droplet-in-oil manner, eliminating the need for chamber sealing. The combination of these design features greatly reduces the complexity in implementing fully integrated lab-on-a-chip systems for genetic detection, facilitating parallel analysis of multiple samples or genes on a single microchip. The capability of the device is demonstrated by performing DNA isolation from the human whole blood sample and analyzing the Rsf-1 gene using the TaqMan probe based gene specific PCR assays.

DC electrokinetic particle transport in an L-shaped microchannel.

Electrokinetic transport of particles through an L-shaped microchannel under DC electric fields is theoretically and experimentally investigated. The emphasis is placed on the direct current (DC) dielectrophoretic (DEP) effect arising from the interactions between the induced spatially nonuniform electric field around the corner and the dielectric particles. A transient multiphysics model is developed in an arbitrary Lagrangian-Eulerian (ALE) framework, which comprises the Navier-Stokes equations for the fluid flow and the Laplace equation for the electrical potential. The predictions of the DEP-induced particle trajectory shift in the L-shaped microchannel are in quantitative agreement with the obtained experimental results. Numerical studies also show that the DEP effect can alter the angular velocity and even the direction of the particle's rotation. Further parametric studies suggest that the L-shaped microfluidic channel may be utilized to focus and separate particles by size via the induced DEP effect.

Particle trapping in high-conductivity media with electrothermally enhanced negative dielectrophoresis.

We demonstrate negative dielectrophoresis (DEP) trapping of particles from high-conductivity media using a novel planar microelectrode that allows electrothermal enhancement of DEP traps. DEP force and electrothermal flow motion are investigated using a scaling analysis, numerical simulations, and experiments. Results show that the DEP trapping is enhanced by lateral transport of particles toward the capture zones due to electrothermal flow, whereas DEP trapping occurred only in limited spatial ranges without the flow motion. The electrothermally enhanced DEP will broaden the limit of electrokinetic manipulations in high-conductivity media. By providing patterned trapping zones that can act as target-specific attachment/detection sites, the presented device allows development of biosensor applications for rapid detection of pathogens and other microorganisms within a practical range of buffer conductivity.

Alternating current electrokinetic motion of colloidal particles on interdigitated microelectrodes.

Alternating current (ac) electrokinetic motion of colloidal particles suspended in an aqueous medium and subjected to a spatially nonuniform ac electric field are examined using a simple theoretical model that considers the relative magnitudes of dielectrophoresis, electrophoresis, ac-electroosmosis, and Brownian motion. Dominant electrokinetic forces are explained as a function of the electric field frequency, amplitude, and conductivity of the suspending medium for given material properties and geometry. Parametric experimental validations of the model are conducted utilizing interdigitated microelectrodes with polystyrene and gold particles and Clostridium sporogenes bacterial spores. The theoretical model provides quantitative descriptions of ac electrokinetic transport for the given target species in a wide spectrum of electric field amplitude and frequency and medium conductivity. The presented model can be used as an effective framework for design and optimization of ac electrokinetic devices.