Colorimetric Assay For Detection Research Articles

A novel light-controlled colorimetric detection assay for nitroreductase based on p-aminophenol-catalyzed and NADH-mediated synthesis of silver nanoparticles.

A novel and efficient light-controlled colorimetric assay for the quantification and detection of nitroreductase (NTR) was constructed based on p-aminophenol (pAP)-catalyzed and nicotinamide adenine dinucleotide (NADH)-mediated generation of AgNPs. Due to the hydrolysis of p-nitrophenol by NTR in the presence of NADH, the hydrolysis product can be used as a catalyst to catalyze the reduction of Ag+ by NADH under the light. As the concentration of NTR increases, the value of absorbance at ca. 400 nm (A400) decreases and the color of the solution turns from brown to bright yellow. A linear correlation was obtained between A400 and the NTR concentration in the range from 1-50 μg mL-1 and the limit of detection (LOD) is 0.27 μg mL-1. The detection system does not respond to other common biological molecules due to the specificity of enzymes and the effect of the nitroreductase inhibitor on the NTR activity was also tested. Finally, we applied the assay to determine NTR in human serum samples by spiking different concentrations of NTR with a recovery of 85.2%-92.5%.

A reusable colorimetric assay based on mixed valence state Ce-MOF@Pt nanoparticles for highly sensitive detection of visfatin

In this work, a colorimetric assay for visfatin detection is described. The mixed valence state Ce-MOF (MVCM) modified by platinum nanoparticles (Pt NPs) is used as a novel catalyst. MVCM exhibits excellent intrinsic peroxidase-like catalytic activity due to the spontaneous recycling of the Ce(III)/Ce(IV) system. Pt NPs serve not only as a carrier of the –NH2-modified single strand DNA (S1) but also as a synergistic catalyst of MVCM. The capture probe (S2) attached to the streptavidin-modified magnetic beads (Mag-SA) could combine with the aptamer to form the Mag-SA/S2/aptamer complex. When in the presence of the target visfatin, aptamer specifically combines with the visfatin, which induces the release of Mag-SA/S2 from the Mag-SA/S2/aptamer complex. At this time, the MVCM@Pt/S1 complex connects with the released Mag-SA/S2, which quickly catalyzes the 3,3,3′,3′-tetramethylbenzidine (TMB), leading to a color change. Under optimal conditions, the absorbance increases linearly when the concentration ranges from 1 ng mL−1 to 100 ng mL−1, and the detection limit is as low as 0.11 ng mL−1. Furthermore, Mag-SA/S2 can be reused at least five times by using the uracil-DNA glycosylase (UDG) and an external magnetic field. The proposed method shows satisfying reproducibility, stability, specificity, and sensitivity, and it was successfully applied to detect visfatin in spiked human serum samples. Thus, it has great potential for clinical research, detection, and catalytic applications.

The intrinsic enzyme mimetic activity of platinum oxide for biosensing of glucose

Given the enzymatic properties and the oxidized surface of Pt nanomaterials, we demonstrated the intrinsic oxidase-like and peroxidase-like activities of platinum oxide (PtO2). The surface clean PtO2 nanoparticles with high water dispersibility were synthesized by a simple and green method. X-ray diffraction (XRD) and X-ray photoelectron spectra (XPS) results demonstrated that the prepared PtO2 nanoparticles mainly consisted of Pt (Ⅳ) state without Pt(0) chemical state. The enzymatic activity of PtO2 nanoparticles was verified by catalytic oxidation of several chromogenic substrates. Catalytic mechanism analysis suggested that PtO2 nanopartiles acted as peroxidase and oxidase enzyme mimics by promoting the generation of the reactive oxygen species (ROS). By combining glucose oxidase, a colorimetric assay for glucose detection was developed with the limit of detection of 10.8 μM. The successful application of the proposed detection assay in human serum samples demonstrated the promising practical application in clinical diagnosis, pharmacy and food.

A rapid colorimetric LAMP assay for detection of Rhizoctonia solani AG-1 IA causing sheath blight of rice

Rhizoctonia solani is one of the most devastating pathogens. R. solani AG-1 IA causes sheath blight in rice, maize, and other Gramineous plants. Accurate identification is essential for the effective management of this pathogen. In the present study, a set of four primers were designed viz. RSPG1, RSPG2, RSPG4, and RSPG5 for polygalacturonase (PG) gene, an important virulence factor in phytopathogenic fungi. All four primer sets showed specific amplification of 300 bp (RSPG1F/R), 375 bp (RSPG2F/R), 500 bp (RSPG4F/R) and 336 bp (RSPG5F/R) amplicons. q-PCR detection using each primer sets could detect up to 10 pg of DNA. We also designed six primers (RS_pg_F3_1/RS_pg_B3_1, RS_pg_FIP_1.1/RS-pg_BIP_1.1, and RS_pg_LF_1/RS_pg_LB_1) for PG gene. Further, a colorimetric LAMP assay developed yielded visual confirmation of the pathogen within 45 min of sample collection when coupled with rapid high throughput template preparation method (rHTTP) from infected samples. The sensitivity of the LAMP assay was as low as 1.65 fg/µl of template DNA and could effectively detect R. solani AG-1 IA from diseased plant tissues and soil samples. The LAMP assay was highly specific for R. solani as it did not show any amplification with other AG groups of R. solani and closely related fungal and bacterial outgroups. This study will help in designing an effective point of care diagnostic method for early monitoring of R. solani and thereby planning timely preventive measures against the pathogen.

ATR-FTIR spectroscopy in blood plasma combined with multivariate analysis to detect HIV infection in pregnant women

The primary concern for HIV-infected pregnant women is the vertical transmission that can occur during pregnancy, in the intrauterine period, during labour or even breastfeeding. The risk of vertical transmission can be reduced by early diagnosis. Therefore, it is necessary to develop new methods to detect this virus in a quick and low-cost fashion, as colorimetric assays for HIV detection tend to be laborious and costly. Herein, attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy combined with multivariate analysis was employed to distinguish HIV-infected patients from healthy uninfected controls in a total of 120 blood plasma samples. The best sensitivity (83%) and specificity (92%) values were obtained using the genetic algorithm with linear discriminant analysis (GA-LDA). These good classification results in addition to the potential for high analytical frequency, the low cost and reagent-free nature of this method demonstrate its potential as an alternative tool for HIV screening during pregnancy.

Paper-based antibiotic sensor (PAS) relying on colorimetric indirect competitive enzyme-linked immunosorbent assay for quantitative tetracycline and chloramphenicol detection

Antibiotics such as tetracycline (TET) and chloramphenicol (CAP) are widely used, but their abuse leads to antibiotic residue which is harmful to human health. To obtain sensitive and accurate TET and CAP detections, we design the paper-based antibiotic sensor (PAS) relying on the colorimetric indirect competitive ELISA. PAS can be simply fabricated only via wax printing and surface modification, making the cost of single detection rather cheap as ∼0.15 US dollar; and additionally, it can be fabricated and conserved in good stability. Moreover, the PAS supporting smartphone application with the functions of image recording, correction and analysis is also designed for quantitative antibiotic detections. Using a series of standard solutions, it is proved that the PAS could reach the extremely low limits of detection as 0.5 ng/mL and 0.05 ng/mL as well as the wide effective linear detecting ranges as 1∼103 ng/mL and 0.1∼100 ng/mL in the TET and CAP detections, respectively. Moreover, using milk and fish samples, it was also verified that the PAS could quantify the TET and CAP concentrations accurately even in the practical applications.

Direct colorimetric LAMP assay for rapid detection of African swine fever virus: A validation study during an outbreak in Vietnam

African swine fever (ASF) is a highly infectious viral disease with high mortality. The most recent ASF outbreak in Vietnam began in 2019, posing a threat to spread to the neighbouring Asian countries. Without a commercial vaccine or efficient chemotherapeutics, rapid diagnosis and necessary biosecurity procedures are required to control the disease. While the diagnostic method of ASF recommended by the World Organization of Animal Health is real-time PCR, the ideal diagnosis procedure including master mix setup, template extraction and a high-cost qPCR equipment for many samples being tested simultaneously is not portable. In this study, a colorimetric loop-mediated isothermal amplification (LAMP) assay was modified and evaluated for ASF virus detection using crude serum samples collected from domestic pigs in Vietnam during the 2019 outbreak. The LAMP results can be readily visualized to the naked eye within 30min without the requirement of DNA extraction and sophisticated equipment. The sensitivity, specificity and limit of detection of direct colorimetric LAMP assay were comparable to a commercial diagnostic real-time PCR kit. Results strongly indicate that the adapted colorimetric LAMP assay has a remarkable potential for the in-field diagnosis of ASF.

Dual-Product Synergistically Enhanced Colorimetric Assay for Sensitive Detection of Lipid Transferase Activity.

Lipid transferase-catalyzed protein lipidation plays critical roles in many physiological processes and it has been an increasingly attractive therapeutic target from cancer to neurodegeneration, while sensitive detection of lipid transferase activity in biological samples remains challenging. Here, we presented an AuNP-based colorimetric method with dual-product synergistically enhanced sensitivity for convenient detection of lipid transferase activity. Homo sapiens N-myristoyltransferase 1 (HsNMT1), a key lipid transferase, was selected as the model. Accordingly, positively charged substrate peptides (Pep) of HsNMT1 can induce the aggregation of AuNPs through disrupting their electrostatic repulsion, while the HsNMT1-catalyzed lipid modification generates aggregated lipidated peptides (C14-Pep) and negatively charged HS-CoA, which will eliminate the disruption and stabilize the AuNPs by the formation of Au-S bonds, respectively. Consequently, charge reversal of the biomolecules and the formation of Au-S bonds synergistically contribute to the stability of AuNPs in the presence of HsNMT1. Therefore, the HsNMT1 activity can be visually detected by the naked eye through the color change of the AuNPs originated from the change in their distance-dependent surface plasmon resonance absorptions. Here, the A520/A610 ratio can sensitively reflect the activity of HsNMT1 in the linear range of 2-75 nM with a low detection limit of 0.56 nM. Moreover, the method was successfully applied for probing the HsNMT1 activities in different cell lysates and inhibitor screening. Furthermore, given the replaceability of the substrate peptide, the proposed assay is promising for universal application to other lipid transferases and exhibits great potential in lipid transferase-targeted drug development.

Breaking the pH limitation of peroxidase-like CoFe2O4 nanozyme via vitriolization for one-step glucose detection at physiological pH

In recent years, a growing number of nanosized materials that can mimic the catalytic characteristics of some natural enzymes have experienced a period of rapid development. Among these nanomaterials defined as ‘nanozymes’, peroxidase-mimicking materials draw intensive interest because of their potential applications in various fields, including biosensing, medicine, and environmental engineering. Nonetheless, most of currently explored peroxidase nanozymes exhibit favorable activity only in acidic environments, which greatly restricts their use especially in biochemical analysis. Developing peroxidase mimics with good catalytic performance in a broad pH range is highly desired and still challenging. Herein, we report a vitriolization strategy to broaden the catalytic pH scope of peroxidase-like CoFe2O4. The vitriolization process endows the nanomaterial with sustainable acidity on its surface. As a result, the obtained peroxidase nanozyme (SO42−/CoFe2O4) exhibits good catalytic activity in neutral or even weak alkaline solutions. This provides us a one-step assay for colorimetric glucose detection in one pot at physiological pH via combining the proposed peroxidase mimic with glucose oxidase, which shows advantages of easier operation and faster detection while offering comparable analytical performance.

Designing a two-stage colorimetric sensing strategy based on citrate reduced gold nanoparticles: Sequential detection of Sanguinarine (anticancer drug) and visual sensing of DNA.

Distance dependent optical properties of colloidal gold nanoparticles offer designing of colorimetric sensing modalities for detection of a variety of analytes. Herein, we report a simple and facile colorimetric detection assay for an anti-cancer drug, Sanguinarine (SNG) and Calf Thymus DNA (Ct-DNA) based on citrate reduced gold nanoparticles (CI-Au NPs). The electrostatic interaction between SNG and CI-Au NPs induce aggregation of Au NPs accompanied with visible colour change of colloidal solution. The assay conditions like salt concentration, pH and reaction time had been adjusted to achieve highly sensitive and fast colorimetric response. Furthermore, the optimized CI-Au NPs/SNG sensing system is used for the detection of Ct-DNA based on the mechanism of anti-aggregation of CI-Au NPs. The simultaneous presence of SNG and Ct-DNA prevent aggregation of Au NPs owing to preferential formation of Ct-DNA-SNG intercalation complex and colour of the Au NPs solution tends to remain red, depending on the concentration of Ct-DNA in solution. The degree of aggregation and anti-aggregation of CI-Au NPs was monitored using Transmission electron microscopic (TEM) measurements and UV-Visible spectrophotometry by analysing the ratio of absorptions for aggregated and dispersed Au NPs. The intercalation mode of binding between SNG and Ct-DNA in CI-Au NPs/SNG sensing system was determined by Fluorescence spectral studies and UV-thermal melting studies. The absorption ratio (A627/A525) of Au NPs exhibited a linear correlation with SNG concentrations in the range from 0 to 0.9μM with detection limit as 0.046μM. This optical method can determine Ct-DNA as low as 0.36μM and the calibration is linear for concentration range 0 to 5μM. The proposed sensing strategy enables detection as well as quantification of SNG & Ct-DNA in real samples with satisfactory results and finds application in drug or DNA monitoring.

Paper chip-based colorimetric assay for detection of Salmonella typhimurium by combining aptamer-modified Fe3O4@Ag nanoprobes and urease activity inhibition.

Arapid and sensitive colorimetric assay is described forSalmonella typhimurium (S. typhimurium) detection using urea/phenol red impregnated test paper. Aptamer-modified Fe3O4@Ag multifunctional hybrid nanoprobes (apt-Fe3O4@Ag NPs) were used to specifically captured S. typhimurium; the nanoprobes were quickly etched by H2O2 to form Ag+. The generated Ag+ can inhibit the urease-catalyzed hydrolysis reaction of urea to produce NH4+. Consequently, the as-prepared test paper displayed a yellow color. In the presence ofS. typhimurium, the target bacteria can cause aggregation of apt-Fe3O4@Ag NPs, and the deposited Ag on the nanoprobe's surface is shielded against H2O2-induced oxidative decomposition leading to reduced Ag+ production. The catalytic activity of urease cannot be inhibited completely by inadequate amount of Ag+. An obvious color change from yellow to pink can be monitored directly using our test paper as a result of increased NH4+. The entire assay procedure could be completed within 1h. A limit of detection of 48cfu/mL is achieved with a linear range of 1×102 to 1×106cfu/mL. The recoveries ofS. typhimurium spiked in pure milk samples were 92.48-94.05%. Graphical abstract Schematic diagram of the proposed colorimetric assay for S. typhimurium detection based on etching of bifunctional apt-Fe3O4@Ag NPs and inhibiting catalytic activity of urease by Ag+. A color change from yellow to pink can be observed and correlated to the concentration of S. typhimurium.

Development of Real-Time and Colorimetric Loop Mediated Isothermal Amplification Assay for Detection of Xanthomonas gardneri

Xanthomonas gardneri is one of the causal agents of bacterial spot (BS), an economically important bacterial disease of tomato and pepper. Field-deployable and portable loop-mediated isothermal amplification (LAMP)-based instruments provide rapid and sensitive detection of plant pathogens. In order to rapidly and accurately identify and differentiate X. gardneri from other BS-causing Xanthomonas spp., we optimized a new real-time monitoring LAMP-based method targeting the X. gardneri-specific hrpB gene. Specificity and sensitivity of real-time and colorimetric LAMP assays were tested on the complex of bacterial strains pathogenic to tomato and pepper and on plants infected by the pathogen. The assay detection limit was 1 pg/μL of genomic DNA with an assay duration of only 30 min. The use of portable and handheld instruments allows for fast analysis, reducing the diagnosis time, and can contribute to proper disease management and control of X. gardneri. Due to the high efficiency of this method, we suggest its use as a standard diagnostic tool during phytosanitary controls.

A multicolor colorimetric assay for sensitive detection of sulfide ions based on anti-etching of triangular gold nanoplates

This study presents a viable colorimetric assay for sensitive sensing of sulfide ions (S2−) using triangular gold nanoplates (AuNPLs) as nanoprobes. The sensing mechanism is based on regulating the etching process of AuNPLs upon the addition of different concentration of S2−. In the presence of Cu2+ and I−, the AuNPLs would be etched by triiodide ions (I3−) produced from the redox reaction between Cu2+ and I−, resulting in the transformation of the shape of AuNPLs from triangular into sphere and blue shifted localized surface plasmon resonance (LSPR). However, in the presence of S2−, a preferential reaction between Cu2+ and S2− occurred, and the inhibition of the formation of I3− prevented the AuNPLs from being etched. Correspondingly, a red shift of LSPR can be observed with an increase of the concentration of S2−, accompanying a readily distinguishable color transition that goes from pink to purplish red, purple, bluish purple and blue. The color alterations allow for a visual estimation of the concentrations of S2−, and the lowest visual concentration through naked-eyes was 0.1 μM. A good linear relationship (R2 = 0.997) between the peak shifts and the concentration of S2− was obtained in the range of 0.02–1.5 μM, and the limit of detection (3 s/k) by UV–vis spectroscopy was as low as 16 nM. The as-established assay displayed excellent selectivity, which was successfully applied for analysis of S2− in environmental water samples.

A novel colorimetric assay for calcium ion and calmodulin detection based on gold nanoparticles

A facile and reliable colorimetric method for detection of calcium ion and calmodulin was developed, which is based on the calcium ion-induced aggregation of gold nanoparticles, accompanied by an obvious color change from red to blue-purple. Calcium ion can be detected in the concentration range from 2.5 to 37.5 µmol·L−1 with a 0.6 µmol·L−1 detection limit. Calmodulin can preferentially capture calcium ion to inhibit the aggregation and the color changes of gold nanoparticles, and can be determined with a 0.8 µg·mL−1 detection limit over the concentration range from 0 to 33.6 µg·mL−1.

A Rapid, Simple, Inexpensive, and Mobile Colorimetric Assay COVID-19-LAMP for Mass On-Site Screening of COVID-19.

To control the COVID-19 pandemic and prevent its resurgence in areas preparing for a return of economic activities, a method for a rapid, simple, and inexpensive point-of-care diagnosis and mass screening is urgently needed. We developed and evaluated a one-step colorimetric reverse-transcriptional loop-mediated isothermal amplification assay (COVID-19-LAMP) for detection of SARS-CoV-2, using SARS-CoV-2 isolate and respiratory samples from patients with COVID-19 (n = 223) and other respiratory virus infections (n = 143). The assay involves simple equipment and techniques and low cost, without the need for expensive qPCR machines, and the result, indicated by color change, is easily interpreted by naked eyes. COVID-19-LAMP can detect SARS-CoV-2 RNA with detection limit of 42 copies/reaction. Of 223 respiratory samples positive for SARS-CoV-2 by qRT-PCR, 212 and 219 were positive by COVID-19-LAMP at 60 and 90 min (sensitivities of 95.07% and 98.21%) respectively, with the highest sensitivities among nasopharyngeal swabs (96.88% and 98.96%), compared to sputum/deep throat saliva samples (94.03% and 97.02%), and throat swab samples (93.33% and 98.33%). None of the 143 samples with other respiratory viruses were positive by COVID-19-LAMP, showing 100% specificity. Samples with higher viral load showed shorter detection time, some as early as 30 min. This inexpensive, highly sensitive and specific COVID-19-LAMP assay can be useful for rapid deployment as mobile diagnostic units to resource-limiting areas for point-of-care diagnosis, and for unlimited high-throughput mass screening at borders to reduce cross-regional transmission.

A Colorimetric Assay to Enable High-Throughput Identification of Biofilm Exopolysaccharide-Hydrolyzing Enzymes.

Glycosidase enzymes that hydrolyze the biofilm exopolysaccharide poly-β-(1→6)-N-acetylglucosamine (PNAG) are critical tools to study biofilm and potential therapeutic biofilm dispersal agents. Function-driven metagenomic screening is a powerful approach for the discovery of new glycosidase but requires sensitive assays capable of distinguishing between the desired enzyme and functionally related enzymes. Herein, we report the synthesis of a colorimetric PNAG disaccharide analogue whose hydrolysis by PNAG glycosidases results in production of para-nitroaniline that can be continuously monitored at 410 nm. The assay is specific for enzymes capable of hydrolyzing PNAG and not related β-hexosaminidase enzymes with alternative glycosidic linkage specificities. This analogue enabled development of a continuous colorimetric assay for detection of PNAG hydrolyzing enzyme activity in crude E. coli cell lysates and suggests that this disaccharide probe will be critical for establishing the functional screening of metagenomic DNA libraries.

A field-applicable colorimetric assay for notorious explosive triacetone triperoxide through nanozyme-catalyzed irreversible oxidation of 3, 3'-diaminobenzidine.

A field-applicable colorimetric assay for fast detection of notorious explosive triacetone triperoxide (TATP) has been developed through the selective irreversible oxidation of 3, 3'-diaminobenzidine by hydrogen peroxide (HP) liberated during the acidic hydrolysis/degradation of TATP in the presence of MnO2 nanozymes. The generated HP was detected by probing the absorbance of the product (indamine polymer) of the 3, 3'-diaminobenzidine (DAB) oxidation reaction at 460.0nm. The UV-Vis measurements provided a linear range from 1.57 to 10.50mgL-1 TATP with a detection limit of 0.34mgL-1. The oxidation of DAB cannot proceed by molecular oxygen, thus it is selectively oxidized by H2O2; this prevents false-positive resultsfrom laundry detergents (containing O2-releasing substances). Moreover, a naked-eye field test was developed, and a fast spot test analyzing time of 5s was achieved. The selectivity of the assay was checked by analyzing some synthetic samples prepared with a laundry detergent as camouflage. The results of the developed assay revealed quantitative recoveries for TATPwhereas the standard nanozyme-based method showed significant false-positiveresults. Graphical abstract.

Water safety screening via multiplex LAMP-Au-nanoprobe integrated approach

Contaminated water resources remain a major global concern regarding public health. The majority of water safety protocols include indicators of microbial contamination to evaluate the potential risk to public health and are key elements of quality guidelines. Among these, markers for total coliforms and fecal coliforms are strong indicators of co-contamination with other pathogens. Traditional methods, recurring to slow and cumbersome culture-based approaches, have been gradually replaced by molecular methods, capable of faster and more specific screening. These are usually PCR-based methods that may allow for multiple pathogen detection but require dedicated laboratory equipment, hindering the rapid on-site assessment.Here, we used a multiplex Loop-Mediated Isothermal Amplification (mLAMP) strategy for the amplification of two markers associated with the contamination by total and fecal coliforms (e.g. Escherichia coli) — lacZ and uidA genes, respectively — thus allowing for single tube multiplex detection. The mLAMP products were then subject to an Au-nanoprobe colorimetric detection assay for precise discrimination of targets. This approach was validated in 22 water samples that were also screened for the presence of lacZ and uidA using standard and quantitative PCR, with the capability for discriminating the contamination level, e.g. a semi-quantitative evaluation of water quality.

Colorimetric Urinalysis for On-Site Detection of Metabolic Biomarkers.

Over the past few decades, colorimetric assays have been developed for cost-effective and rapid on-site urinalysis. Most of these assays were employed for detection of biomarkers such as glucose, uric acid, ions, and albumin that are abundant in urine at micromolar to millimolar levels. In contrast, direct assaying of urinary biomarkers such as glycated proteins, low-molecular-weight reactive oxygen species, and nucleic acids that are present at significantly lower levels (nanomolar to picomolar) remain challenging due to the interferences from the urine sample matrix. State-of-the-art assays for detection of trace amounts of urinary biomarkers typically utilize time-consuming and equipment-dependent sample pretreatment or clean-up protocols prior to assaying, which limits their applicability for on-site analysis. Herein, we report a colorimetric assay for on-site detection of trace amount of generic biomarkers in urine without involving tedious sample pretreatment protocols. The detection strategy is based on monitoring the changes in optical properties of poly(3-(4-methyl-3'-thienyloxy)propyltriethylammonium bromide) upon interacting with an aptamer or a peptide nucleic acid in the presence and absence of target biomarkers of relevance for the diagnosis of metabolic complications and diabetes. As a proof of concept, this study demonstrates facile assaying of advanced glycation end products, 8-hydroxy-2'-deoxyguanosine and hepatitis B virus DNA in urine samples at clinically relevant concentrations, with limits of detection of ∼850 pM, ∼650 pM, and ∼ 1 nM, respectively. These analytes represent three distinct classes of biomarkers: (i) glycated proteins, (ii) low-molecular-weight reactive oxygen species, and (iii) nucleic acids. Hence, the proposed methodology is applicable for rapid detection of generic biomarkers in urine, without involving sophisticated equipment and skilled personnel, thereby enabling on-site urinalysis. At the end of the contribution, we discuss the opportunity to translate the homogeneous assay into a paper-based format.

Cas12a-Activated Universal Field-Deployable Detectors for Bacterial Diagnostics

Field-deployable detectors of disease biomarkers provide a simple and fast analysis of clinical specimens. However, most of the existing field-deployable diagnostics have poor sensitivity and are not suitable for the detection of biomarkers with low abundance. Herein, we report a highly sensitive and rapid colorimetric readout paper-based assay for pathogen detection by integrating the unique collateral activity of a Cas12a-activated universal field-deployable detector (CUFD). The collateral effect of Cas12a results in a nonspecific destruction of a fluorophore biotin-labeled ssDNA reporter for the CUFD. This technique can quantify seven different kinds of pathogens in blood samples without any purification procedure, with sensitivity as low as 10 aM for the Shigella dysenteriae DNA. This CUFD technique has significant potential for the detection of pathogenic DNA as well as other types of DNA or RNA targets at the point-of-care application.