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Tuberculosis (TB), one of the deadliest infectious diseases, is caused by Mycobacterium tuberculosis (MTB) and remains a public health problem nowadays. Conventional MTB DNA detection methods require sophisticated infrastructure and well-trained personnel, which leads to increasing complexity and high cost for diagnostics and limits their wide accessibility in low-resource settings. To address these issues, we have developed a low-cost photothermal biosensing method for the quantitative genetic detection of pathogens such as MTB DNA on a paper hybrid device using a thermometer. First, DNA capture probes were simply immobilized on paper through a one-step surface modification process. After DNA sandwich hybridization, oligonucleotide-functionalized gold nanoparticles (AuNPs) were introduced on paper and then catalyzed the oxidation reaction of 3,3',5,5'-tetramethylbenzidine (TMB). The produced oxidized TMB, acting as a strong photothermal agent, was used for the photothermal biosensing of MTB DNA under 808 nm laser irradiation. Under optimal conditions, the on-chip quantitative detection of the target DNA was readily achieved using an inexpensive thermometer as a signal recorder. This method does not require any expensive analytical instrumentation but can achieve higher sensitivity and there are no color interference issues, compared to conventional colorimetric methods. The method was further validated by detecting genomic DNA with high specificity. To the best of our knowledge, this is the first photothermal biosensing strategy for quantitative nucleic acid analysis on microfluidics using a thermometer, which brings fresh inspirations on the development of simple, low-cost, and miniaturized photothermal diagnostic platforms for quantitative detection of a variety of diseases at the point of care.

Conventional paper lateral flow assays have low sensitivity and suffer from severe interference from complex human fluid sample matrices, which prevents their practical application in the testing of whole blood samples in the point-of-care settings. To solve this problem, gold nanostar@Raman reporter@silica-sandwiched nanoparticles have been developed as the surface-enhanced Raman scattering (SERS) probes for sensing transduction; and a functionalized filter membrane assembly has been designed and constructed in the paper-based lateral flow strip (PLFS) as a built-in plasma separation unit. In this "on-strip" plasma separation unit, three layers of filter membranes are stacked and surface-modified to maximize the separation efficiency and the plasma yield. As a result, the integrated PLFS has been successfully used for the detection of carcinoembryonic antigen (CEA) in 30 μL of whole blood with the assistance of a portable Raman reader, achieving a limit of detection of 1.0 ng mL-1. In short, this report presents an inexpensive, disposable, portable, and field-deployable paper-based device as a general point-of-care testing tool for protein biomarker detection in a drop of whole blood.

The analysis of protein antigens as biomarkers in clinical samples is particularly helpful for the early diagnosis of diseases. However, this is difficult to accomplish owing to the presence of the antigens in trace amounts as well as the complexity of the matrixes in clinical samples. In this study, a lab-on-membrane platform that can be combined with paper spray ionization mass spectrometry was developed for the in situ high-throughput sensitive detection of the prostate-specific antigen (PSA). The sensitivity of the proposed platform was enhanced via two strategies: (1) the synthesis of a biotin-streptavidin scaffold caused an increase in the capturing efficiency of PSA by a factor of 5 and (2) the immobilization of a large number of mass tag molecules on the gold nanoparticles allowed for the amplification of the mass spectrometry signals. The limit of detection was approximately 3.0 pg mL-1. The selectivity to PSA was guaranteed by using an antibody-aptamer pairing sandwich immunoassay, and PSA detection was unaffected even when other protein antigens (carcinoembryonic antigen and carbohydrate antigen 125) were present. The modified membranes maintained their performance for at least 30 days when stored at 4 °C. Finally, analysis of human serum samples confirmed that the PSA concentration as determined using the proposed platform was consistent with that determined with a conventional chemiluminescent immunoassay. Thus, this PSA analyzing platform is suitable for prostate cancer diagnosis in clinical settings.

Porous media made of nitrocellulose and glass fiber are common "paper" substrates for lateral flow assays, microfluidic paper analytical devices and other point-of-care diagnostic assays. Such assays commonly use optical labels such as gold nanoparticles, latex beads, or fluorescent nanoparticles to visualize the presence of analytes. Fluorescent labels are commonly used in bioassays to enhance sensitivity, but autoluminescence of the paper substrate worsens signal-to-noise ratios of fluorescence-based assays. To date, there exists no systematic investigation of autoluminescence wavelengths or lifetimes of porous membranes used in lateral flow assays. In response, we quantified the autoluminescence of commonly used porous materials across the visible spectrum via excitation-emission spectroscopy and time-resolved fluorescence spectroscopy, and demonstrate that autoluminescence is solely due to autofluorescence with lifetimes of about 5 ns in the visible spectrum. Counterintuitively, we found that spectroscopy alone does not provide sufficient information to select candidate paper substrates for fluorophore-labeled assays. Therefore, we developed a simple quantitative framework to select a low-fluorescence substrate that minimizes both the overlap of paper and fluorophore emission spectra and the fluorescence intensity on an imaging system of interest (such as a gel imager). Use of this framework was shown to lower the limit of detection of an influenza A nucleoprotein immunoassay by over 50%. The tools developed in this manuscript enable assay developers to screen appropriate, low-fluorescence porous substrates and enhance the sensitivity of membrane-based fluorescence assays.

The development of highly sensitive and selective methods for the detection of microRNA (miRNA) has attracted tremendous attention because of its importance in fundamental biological studies and diagnostic applications. In this work, we develop DNA-encoded Raman-active anisotropic nanoparticles modified origami paper analytical devices (oPADs) for rapid, highly sensitive, and specific miRNA detection. The Raman-active anisotropic nanoparticles were prepared using 10-mer oligo-A, -T, -C, and -G to mediate the growth of Ag cubic seeds into Ag nanoparticles (AgNPs) with different morphologies. The resulting AgNPs were further encoded with DNA probes to serve as effective surface-enhanced Raman scattering (SERS) probes. The analytical device was then fabricated on a single piece of SERS probes loaded paper-based substrate and assembled based on the principles of origami. The addition of the target analyte amplifies the Raman signals on DNA-encoded AgNPs through a target-dependent, sequence specific DNA hybridization assembly. This simple and low-cost analytical device is generic and applicable to a variety of miRNAs, allowing detection sensitivity down to 1 pM and assay time within 15 min, and therefore holds promising applications in point-of-care diagnostics.

Integrating ratiometric photoelectrochemical (PEC) techniques with paper microfluidics to construct a ratiometric PEC paper analytical device for practical application is often restricted by the grave dependence of ratiometric assay on photoactive materials and low mass-transfer rates of the paper channel. Herein, a universal donor/acceptor-induced ratiometric PEC paper analytical device with a hollow double-hydrophilic-walls channel (HDHC) was fabricated for high-performance microRNA-141 (miRNA-141) quantification. Concretely, a photoanode and photocathode were integrated on the paper-based sensing platform in which the photocathode served as a biosensing site for the pursuit of higher selectivity. For formulation of a cascading signal amplification strategy, a unique duplex-specific nuclease-induced target recycling reaction was engineered for the output of a double amount of all useful DNA linkers instead of conventional output of only one available DNA product, which could guarantee the output of abundant DNA linkers with the initiation of a cascade of hybridization chain reaction on both the trunk and branch in the presence of miRNA-141. Then the formed dendriform polymeric DNA duplex structures were further decorated with glucose oxidase (GOx)-mimicking gold nanoparticles by the electrostatic interaction to form a branchy gold tree (BGT). Profiting from the perfect GOx-mimicking activity of BGT and high mass-transfer rates of HDHC, the cathodic photocurrent from Ag

To achieve personalized healthcare, a quick, accurate, and high-throughput method to detect disease biomarkers is essential. In the traditional practice, mass spectrometry is one of the most powerful tools and is widely studied. However, the test of human serum usually requires complicated sample pretreatment, tedious operations, and precise condition control, especially for the detection of enzymes as biomarkers. As butyrylcholinesterase (BuChE) has an indicative significance in detecting degenerative disease, liver injury, and organophosphate poisoning, the quick quantification of BuChE is of vital importance to the clinic. In this paper, we report the design and fabrication of a portable 3D-printed enzyme reactor paper spray cartridge (3D ER-PS) with integrated functions: temperature control, enzyme reaction, analyte transfer, and paper spray ionization. Coupled with mass spectrometry, quantitative testing of BuChE activity in human serum was realized conveniently and accurately. While it only requires very simple sample preparation, the results from current 3D ER-PS approach are well consistent with those obtained using Ellman's method. This 3D ER-PS platform not only provides a novel solution for the liquid biopsy of BuChE in clinics but also contributes to the development of quick and targeted medical approaches for analyzing other types of serum biomarker molecules in the field of disease diagnosis.

Paper-based lateral flow immunoassays (LFIAs) using conventional sandwich-type immunoassays are one of the most commonly used point-of-care (PoC) tests. However, the application of gold nanoparticles (AuNPs) in LFIAs does not meet sensitivity requirements for the detection of infectious diseases or biomarkers present at low concentrations in body fluids because of the limited number of AuNPs that can bind to the target. To overcome this problem, we first developed a single-stranded DNA binding protein (RPA70A, DNA binding domain A of human Replication Protein A 70 kDa) conjugated to AuNPs for a sandwich assay using a capture antibody immobilized in the LFIA and an aptamer as a detection probe, thus, enabling signal intensity enhancement by attaching several AuNPs per aptamer. We applied this method to detect the influenza nucleoprotein (NP) and cardiac troponin I (cTnI). We visually detected spiked targets at a low femtomolar range, with limits of detection for NP in human nasal fluid and for cTnI in serum of 0.26 and 0.23 pg·mL-1, respectively. This technique showed significantly higher sensitivity than conventional methods that are widely used in LFIAs involving antibody-conjugated AuNPs. These results suggest that the proposed method can be universally applied to the detection of substances requiring high sensitivity and can be used in the field of PoC testing for early disease diagnosis.

In this work, a novel dual photoelectrochemical/colorimetric cyto-analysis format was first introduced into a microfluidic paper-based analytical device (μ-PAD) for synchronous sensitive and visual detection of H2O2 released from tumor cells based on an in situ hydroxyl radicals ((•)OH) cleaving DNA approach. The resulted μ-PAD offered an excellent platform for high-performance biosensing applications, which was constructed by a layer-by-layer modification of concanavalin A, graphene quantum dots (GQDs) labeled flower-like Au@Pd alloy nanoparticles (NPs) probe, and tumor cells on the surface of the vertically aligned bamboo like ZnO, which grows on a pyknotic Pt NPs modified paper working electrode (ZnO/Pt-PWE). It was the effective matching of energy levels between GQDs and ZnO levels that lead to the enhancement of the photocurrent response compared with the bare ZnO/Pt-PWE. After releasing H2O2, the DNA strand was cleaved by (•)OH generated under the synergistic catalysis of GQDs and Au@Pd alloy NPs and thus, reduced the photocurrent, resulting in a high sensitivity to H2O2 in aqueous solutions with a detection limit of 0.05 nmol observed, much lower than that in the previously reported method. The disengaged probe can result in catalytic chromogenic reaction of substrates, resulting in real-time imaging of H2O2 biological processes. Therefore, this work provided a truly low-cost, simple, and disposable μ-PAD for precise and visual detection of cellular H2O2, which had potential utility to cellular biology and pathophysiology.

In nucleic acid testing (NAT), gold nanoparticle (AuNP)-based lateral flow assays (LFAs) have received significant attention due to their cost-effectiveness, rapidity, and the ability to produce a simple colorimetric readout. However, the poor sensitivity of AuNP-based LFAs limits its widespread applications. Even though various efforts have been made to improve the assay sensitivity, most methods are inappropriate for integration into LFA for sample-to-answer NAT at the point-of-care (POC), usually due to the complicated fabrication processes or incompatible chemicals used. To address this, we propose a novel strategy of integrating a simple fluidic control strategy into LFA. The strategy involves incorporating a piece of paper-based shunt and a polydimethylsiloxane (PDMS) barrier to the strip to achieve optimum fluidic delays for LFA signal enhancement, resulting in 10-fold signal enhancement over unmodified LFA. The phenomena of fluidic delay were also evaluated by mathematical simulation, through which we found the movement of fluid throughout the shunt and the tortuosity effects in the presence of PDMS barrier, which significantly affect the detection sensitivity. To demonstrate the potential of integrating this strategy into a LFA with sample-in-answer-out capability, we further applied this strategy into our prototype sample-to-answer LFA to sensitively detect the Hepatitis B virus (HBV) in clinical blood samples. The proposed strategy offers great potential for highly sensitive detection of various targets for wide application in the near future.

We report the first portable upconversion nanoparticles (UCNPs)-based paper device for road-side field testing of cocaine. Upon the recognition of cocaine by two pieces of rationally designed aptamer fragments, the luminescence of UCNPs immobilized on the paper is quenched by Au nanoparticles (AuNPs), which indicates the cocaine concentration. This device can give quantitative results in a short time with high sensitivity using only a smartphone as the apparatus. Moreover, this device is applicable in human saliva samples, and it also can be used to monitor the cocaine content change in blood samples. The results of this work demonstrate the prospect of developing UCNPs-based paper devices for field testing of drug abuse.

A novel paper electrode-based flexible pressure sensor modified with multiwalled carbon nanotubes was designed for point-of-care (POC) immunoassay of carcinoembryonic antigen (CEA) with digital multimeter readout. The portable POC testing device consisted of flexible pressure sensor equipped with a paper electrode and connected through syringe tubing to a single-break microplate. The immunoreaction was initially carried out on the microplate with a sandwich-type assay format using platinum nanozyme-labeled secondary antibody for the gas generation. Upon addition of hydrogen peroxide (H2O2), platinum nanozyme (catalase-like mimic) reduced it into hydrogen oxide and oxygen (O2). The overflowing oxygen gas increased the pressure of the multiwalled carbon nanotube-functionalized paper electrode in a homemade pressure-tight system, and the increased pressure could be readily monitored using the paper electrode-based flexible pressure-tight sensor with a digital multimeter readout. The detectable signal mainly ...

This work has looked to explore an innovative and powerful visible fluorescence immunoassay method through wet NH

The detection of double stranded DNA (dsDNA) is often associated with the use of laboratory-bound approaches and/or with the prior generation of single stranded DNA (ssDNA), making these methods not suitable for in situ monitoring, i.e., point-of-care diagnostics. Screen-printed technology, coupled to the use of triplex forming oligonucleotides (TFO) as the recognizing probes, offers a great possibility toward the development of portable analytical tools. Moreover, the continuous demand for sustainable processes and waste lowering have highlighted the role of paper-based substrates for manufacturing easy-to-use, low-cost, and sustainable electrochemical devices. In this work, filter paper and copy paper have been utilized to produce E-DNA strips. Gold nanoparticles (AuNPs) have been exploited to immobilize the methylene blue (MB)-tagged TFO and to enhance the charge transfer kinetics at the electrode surface. Both paper-based substrates have been electrochemically characterized, and in addition, the effect of the amount of waxed layers has been evaluated. The paper-based E-DNA strips have been challenged toward the detection of three model targets, obtaining 3 and 7 nM as the detection limit, respectively, for single and double stranded sequences. The repeatability of the manufacturing (homemade) process has been evaluated with a relative standard deviation of approximately 10%. The effectiveness of the filter paper-based platform has been also evaluated in undiluted serum obtaining a similar value of the detection limit (compared to the measurements carried out in buffer solution). In addition, a synthetic PCR amplified dsDNA sequence, related to HIV, has been detected in serum samples.

The identification of fentanyl, a main culprit in opioid overdose deaths, has become critical. Whereas Raman spectroscopy is an effective tool for detecting illicit drugs, the weak intensity of Raman scattering can make it difficult to distinguish trace materials. This shortcoming is addressed by surface-enhanced Raman spectroscopy (SERS), which produces strong signal enhancements when target compounds are near metal nanoparticles. This work examines the use of a paper-based substrate impregnated with silver nanoparticles for the detection of trace quantities of fentanyl alone and as an adulterant in heroin. In addition, intensity ratios of diagnostic peaks associated with each substance were fitted to a Langmuir isotherm calibration model and used for the quantitative analysis of fentanyl in heroin mixtures. Linearity was observed at6% fentanyl, a significant finding that is consistent with concentrations found in drugs seized during law enforcement efforts. In addition, swabbing with these paper-based SERS substrates facilitated the recovery of fentanyl from surfaces, showing this to be applicable for crime scene investigations. However, assessment using the calibration model proved difficult for swabbed samples. Overall, this work demonstrates a potentially simple and sensitive technique for the forensic analysis and quantitation of fentanyl in trace amounts.

The development of simple fluorescent and colorimetric assays that enable point-of-care DNA and RNA detection has been a topic of significant research because of the utility of such assays in resource limited settings. The most common motifs utilize hybridization to a complementary detection strand coupled with a sensitive reporter molecule. Here, a paper-based colorimetric assay for DNA detection based on pyrrolidinyl peptide nucleic acid (acpcPNA)-induced nanoparticle aggregation is reported as an alternative to traditional colorimetric approaches. PNA probes are an attractive alternative to DNA and RNA probes because they are chemically and biologically stable, easily synthesized, and hybridize efficiently with the complementary DNA strands. The acpcPNA probe contains a single positive charge from the lysine at C-terminus and causes aggregation of citrate anion-stabilized silver nanoparticles (AgNPs) in the absence of complementary DNA. In the presence of target DNA, formation of the anionic DNA-acpcPNA duplex results in dispersion of the AgNPs as a result of electrostatic repulsion, giving rise to a detectable color change. Factors affecting the sensitivity and selectivity of this assay were investigated, including ionic strength, AgNP concentration, PNA concentration, and DNA strand mismatches. The method was used for screening of synthetic Middle East respiratory syndrome coronavirus (MERS-CoV), Mycobacterium tuberculosis (MTB), and human papillomavirus (HPV) DNA based on a colorimetric paper-based analytical device developed using the aforementioned principle. The oligonucleotide targets were detected by measuring the color change of AgNPs, giving detection limits of 1.53 (MERS-CoV), 1.27 (MTB), and 1.03 nM (HPV). The acpcPNA probe exhibited high selectivity for the complementary oligonucleotides over single-base-mismatch, two-base-mismatch, and noncomplementary DNA targets. The proposed paper-based colorimetric DNA sensor has potential to be an alternative approach for simple, rapid, sensitive, and selective DNA detection.

Though microfluidic paper analytical devices (μPADs) have attracted paramounting attentions in recent years as promising devices for low cost point-of-care tests, their real applications for blood analysis are still challenged by integrating sample preparation with different detection modes on a same μPAD. Herein, we developed a novel μPAD, which well coupled automatic serum extraction with reliable dual mode iron health tests: fluorescent analysis for Fe

We introduce a surface-enhanced Raman scattering (SERS)-functionalized, gold nanoparticle (GNP)-deposited paper strip capable of label-free biofluid sensing for the early detection of infectious eye diseases. The GNP biosensing paper strip was fabricated by the direct synthesis and deposition of GNPs on wax-divided hydrophilic areas of a permeable porous substrate through a facile, power-free synthesizable, and highly reproducible successive ionic layer absorption and reaction (SILAR) technique. To maximize localized surface plasmon resonance-generated SERS activity, the concentration of the reactive solution and number of SILAR cycles were optimized by controlling the size and gap distance of GNPs and verified by computational modeling with geometrical hypotheses of Gaussian-estimated metallic nanoparticles. The responses of our SERS-functionalized GNP paper strip to Raman intensities exhibited an enhancement factor of 7.8 × 10(8), high reproducibility (relative standard deviation of 7.5%), and 1 pM 2-naphthalenethiol highly sensitive detection limit with a correlation coefficient of 0.99, achieved by optimized SILAR conditions including a 10/10 mM/mM HAuCl4/NaBH4 concentration and six SILAR cycles. The SERS-functionalized GNP paper is supported by a multivariate statistics-preprocessed machine learning-judged bioclassification system to provide excellent label-free chemical structure sensitivity for identifying infectious keratoconjunctivitis. The power-free synthesizable fabrication, label-free, rapid analysis, and high sensitivity feature of the SILAR-fabricated SERS-functionalized GNP biosensing paper strip makes it an excellent alternative in point-of-care applications for the early detection of various infectious diseases.

This paper presents a pressed paper-based dipstick that enables detection of foodborne pathogens with multistep reactions by exploiting the delayed fluid flow and channel partition formation on nitrocellulose (NC) membrane. Fluid behaviors are easily modified by controlling the amount of pressure and the position of pressed region on the NC membrane. Detection region of the dipstick is optimized by controlling flow rate and delayed time based on Darcy's law. All the reagents required for assay are dried on the NC membrane and they are sequentially rehydrated at the prepartitioned regions when the device is dipped into sample solution. In this manner, multistep reactions can be facilitated by one-step dipping of the dipstick into the sample solution. As a proof of concept, we performed detection of two fatal foodborne pathogens (e.g., Escherichia coli O157:H7 and Salmonella typhimurium) with signal enhancement. In addition, we expanded the utilization of channel partitions by developing a pressed paper-based dipstick into dual detection format.

Convenient, rapid, and accurate detection of chemical and biomolecules would be a great benefit to medical, pharmaceutical, and environmental sciences. Many chemical and biosensors based on metal nanoparticles (NPs) have been developed. However, as a result of the inconvenience and complexity of most of the current preparation techniques, surface plasmon-based test papers are not as common as, for example, litmus paper, which finds daily use. In this paper, we propose a convenient and practical technique, based on the photothermal effect, to fabricate the plasmonic test paper. This technique is superior to other reported methods for its rapid fabrication time (a few seconds), large-area throughput, selectivity in the positioning of the NPs, and the capability of preparing NP arrays in high density on various paper substrates. In addition to their low cost, portability, flexibility, and biodegradability, plasmonic test paper can be burned after detecting contagious biomolecules, making them safe and eco-friendly.

Here we show that a simple paper-based electrochemical sensor, fabricated by paper folding, is able to detect a 30-base nucleotide sequence characteristic of DNA from the hepatitis B virus (HBV) with a detection limit of 85 pM. This device is based on design principles we have reported previously for detecting proteins via a metalloimmunoassay. It has four desirable attributes. First, its design combines simple origami (paper folding) assembly, the open structure of a hollow-channel paper analytical device to accommodate micrometer-scale particles, and a convenient slip layer for timing incubation steps. Second, two stages of amplification are achieved: silver nanoparticle labels provide a maximum amplification factor of 250 000 and magnetic microbeads, which are mobile solid-phase supports for the capture probes, are concentrated at a detection electrode and provide an additional ∼25-fold amplification. Third, there are no enzymes or antibodies used in the assay, thereby increasing its speed, stability, and robustness. Fourth, only a single sample incubation step is required before detection is initiated.

An on-field colorimetric sensing strategy employing gold nanoparticles (AuNPs) and a paper-based analytical platform was investigated for mercury ion (Hg(2+)) detection at water sources. By utilizing thymine-Hg(2+)-thymine (T-Hg(2+)-T) coordination chemistry, label-free detection oligonucleotide sequences were attached to unmodified gold nanoparticles to provide rapid mercury ion sensing without complicated and time-consuming thiolated or other costly labeled probe preparation processes. Not only is this strategy's sensing mechanism specific toward Hg(2+), rather than other metal ions, but also the conformational change in the detection oligonucleotide sequences introduces different degrees of AuNP aggregation that causes the color of AuNPs to exhibit a mixture variance. To eliminate the use of sophisticated equipment and minimize the power requirement for data analysis and transmission, the color variance of multiple detection results were transferred and concentrated on cellulose-based paper analytical devices, and the data were subsequently transmitted for the readout and storage of results using cloud computing via a smartphone. As a result, a detection limit of 50 nM for Hg(2+) spiked pond and river water could be achieved. Furthermore, multiple tests could be performed simultaneously with a 40 min turnaround time. These results suggest that the proposed platform possesses the capability for sensitive and high-throughput on-site mercury pollution monitoring in resource-constrained settings.

We report a new type of paper analytical device that provides quantitative electrochemical output and detects concentrations as low as 767 fM. The model analyte is labeled with silver nanoparticles (AgNPs), which provide 250 000-fold amplification. AgNPs eliminate the need for enzymatic amplification, thereby improving device stability and response time. The use of magnetic beads to preconcentrate the AgNPs at the detection electrode further improves sensitivity. Response time is improved by incorporation of a hollow channel, which increases the flow rate in the device by a factor of 7 and facilitates the use of magnetic beads. A key reaction necessary for label detection is made possible by the presence of a slip layer, a fluidic switch that can be actuated by manually slipping a piece of paper. The design of the device is versatile and should be useful for detection of proteins, nucleic acids, and microbes.

A simple approach to the mass production of nanoporous gold electrode arrays on cellulose membranes for electrochemical sensing of oxygen using ionic liquid (IL) electrolytes was established. The approach, combining the inkjet printing of gold nanoparticle (GNP) patterns with the self-catalytic growth of these patterns into conducting layers, can fabricate hundreds of self-designed gold arrays on cellulose membranes within several hours using an inexpensive inkjet printer. The resulting paper-based gold electrode arrays (PGEAs) had several unique properties as thin-film sensor platforms, including good conductivity, excellent flexibility, high integration, and low cost. The porous nature of PGEAs also allowed the addition of electrolytes from the back cellulose membrane side and controllably produced large three-phase electrolyte/electrode/gas interfaces at the front electrode side. A novel paper-based solid-state electrochemical oxygen (O(2)) sensor was therefore developed using an IL electrolyte, 1-butyl-3-methylimidazolium hexafluorophosphate (BMIMPF(6)). The sensor looked like a piece of paper but possessed high sensitivity for O(2) in a linear range from 0.054 to 0.177 v/v %, along with a low detection limit of 0.0075% and a short response time of less than 10 s, foreseeing its promising applications in developing cost-effective and environment-friendly paper-based electrochemical gas sensors.

The majority of bioassays utilize thermosensitive reagents (e.g., biomolecules) and laboratory conditions for analysis. The developing world, however, requires inexpensive, simple-to-perform tests that do not require refrigeration or access to highly trained technicians. To address this need, paper-based bioassays using gold nanoparticle (AuNP) colorimetric probes have been developed. In the two prototype DNase I and adenosine-sensing assays, blue (or black)-colored DNA-cross-linked AuNP aggregates were spotted on paper substrates. The addition of target DNase I (or adenosine) solution dissociated the gold aggregates into dispersed AuNPs, which generated an intense red color on paper within one minute. Both hydrophobic and (poly(vinyl alcohol)-coated) hydrophilic paper substrates were suitable for this biosensing platform; by contrast, uncoated hydrophilic paper caused "bleeding" and premature cessation of the assay due to surface drying. The assays are surprisingly thermally stable. During preparation, AuNP aggregate-coated papers can be dried at elevated temperatures (e.g., 90 degrees C) without significant loss of biosensing performance, which suggests the paper substrate protects AuNP aggregate probes from external nonspecific stimuli (e.g., heat). Moreover, the dried AuNP aggregate-coated papers can be stored for at least several weeks without loss of the biosensing function. The combination of paper substrates and AuNP colorimetric probes makes the final products inexpensive, low-volume, portable, disposable, and easy-to-use. We believe this simple, practical bioassay platform will be of interest for use in areas such as disease diagnostics, pathogen detection, and quality monitoring of food and water.