One stone two birds:electrochemical and colorimetric dual-mode biosensor based on copper peroxide/covalent organic framework nanocomposite for ultrasensentive norovirus detection

Norovirus(NoV)is regarded as one of the most common causes of foodborne diarrhea in the world.It is urgent to identify the pathogenic microorganism of the diarrhea in short time.In this work,we developed an electrochemical and colorimetric dual-mode detection for NoV based on the excellent dual catalytic properties of copper peroxide/COF-NH_(2)nanocomposite(CuO_(2)@COF-NH_(2)).For the colorimetric detection,NoV can be directly detected by the naked eye based on CuO_(2)@COF-NH_(2)as a laccase-like nonazyme using“peptide-NoV-antibody”recognition mode.The colorimetric assay displayed a wide and quality linear detection range from 1 copy/mL to 5000 copies/mL of NoV with a low limit of detection(LOD)of 0.125 copy/mL.For the electrochemical detection of NoV,CuO_(2)@COF-NH_(2)showed an oxidation peak of copper ion from Cu^(+)to Cu^(2+)using“peptide-NoV-antibody”recognition mode.The electrochemical assay showed a linear detection range was 1-5000 copies/mL with a LOD of 0.152 copy/mL.It's worthy to note that this assay does not need other electrical signal molecule,which provide the stable and sensitive electrochemial detection for NoV.The electrochemical and colorimetric dual-mode detection was used to detect NoV in foods and faceal samples,which has the potential for improving food safety and diagnosing of NoV-infected diarrhea.

Electrochemical and colorimetric dual-signal detection of Staphylococcus aureus enterotoxin B based on AuPt bimetallic nanoparticles loaded Fe-N-C single atom nanocomposite

Sensitive detection of Staphylococcus aureus enterotoxin B(SEB)is of importance for preventing food poisoning from threatening human health.In this work,an electrochemical and colorimetric dual-signal detection assay of SEB was developed.The probe(Ab2/AuPt@Fe-N-C)was bound to SEB captured by Ab1,where the Ab2/AuPt@Fe-N-C triggered methylene blue degradation and resulted in the decrease of electrochemical signal.Furthermore,the probe catalyzed the oxidation of 3,3’,5,5’-tetramethyl biphenyl to generate a colorimetric absorbance at 652 nm.Once the target was captured and formed a sandwich-like complex,the color changed from colorless to blue.SEB detection by colorimetric and electrochemical methods showed a linear relationship in the concentration ranges of 0.0002-10.0000 and 0.0005-10.0000 ng/mL,with limits of detection of 0.0667 and 0.1670 pg/mL,respectively.The dual-signal biosensor was successfully used to detect SEB in milk and water samples,which has great potential in toxin detection in food and the environment.

Comparison of chemiluminescence enzyme immunoassay based on magnetic microparticles with traditional colorimetric ELISA for the detection of serum α-fetoprotein

A chemiluminescence enzyme immunoassay based on magnetic microparticles (MmPs-CLEIA) was developed to evaluate serum a-fetoprotein (AFP) in parallel with traditional colorimetric enzyme-linked immunosorbent assay (ELISA).A systematic comparison between the MmPs-CLEIA and colorimetric ELISA concluded that the MPs-CLEIA exhibited fewer dosages of immunoreagents,less total assay time,and better linearity,recovery,precision,sensitivity and validity.AFP was detected in forty human serum samples by the proposed MPs-CLEIA and ELISA,and the results were compared with commercial electrochemiluminescence immunoassay (ECLIA) kit.The correlation coefficient between MPs-CLEIA and ELISA was obtained with R 2 0.6703;however,the correlation between MPs-CLEIA and ECLIA (R 2 0.9582) was obviously better than that between colorimetric ELISA and ECLIA (R 2 0.6866).

A rapid and accurate 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide colorimetric assay for quantification of bacteriocins with nisin as an example

The objective of this study is to propose a more accurate and faster MTT 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide colorimetric assay (MCA) for quantitative measurement of polypeptide bacteriocins in solutions with nisin as an example. After an initial incubation of nisin and indicator bacterium Micrococcus luteus NCIB 8166 in tubes, MTT was added for another incubation period. After that, nisin was quantified by estimating the number of viable bacteria based on measuring the amount of purple formazan produced by cleavage of yellow tetrazolium salt MTT. Then MCA was compared to a standard agar diffusion assay (ADA). The results suggested a high correlation coefficient (r2=0.975±0.004) between optical density (OD) and the inhibitory effect of nisin on a bacterial strain Micrococcus luteus NCIB 8166 at a range of 0.125～32 IU/ml. The MCA described in this study was very quick. Quantification of nisin took only 7～8 h and the detection limit was at the level of 0.125 IU/ml when compared to 12 IU/ml and 24～28 h for ADA. The MCA provides an accurate and rapid method for quantifi-cation of nisin in solutions and is expected to be used for quantification of other antimicrobial substances.

Lectin Conjugated Gold Nanoparticle-based Colorimetric Assay for Studying the Interactions of Antibiotic with Living Cell

The interactions of antibiotic with living cells were studied by lectin conjugated gold nanoparticles（GNPs） based colorimetric assay. Because of the high affinity of lectin for saccharides, the lectin conjugated GNPs are able to employ as indicators for monitoring the antibiotic induced changes of glycosyl complexes. The interactions of a well known antibiotic, tunicamycin, with two different cell lines, HeLa and SHG-44, were selected to establish this assay. In the presence of tunicamycin, the dose- and time-dependence on the decreasing of binding affinity of lectin conjugated GNPs with living cells were demonstrated by conventional microscopic and UV-Vis spectroscopic studies. The experimental result demonstrates that our approach can be used to identify antibiotic induced expression difference of glycosyl complexes on different cellular surfaces and determine drug activity quantitatively. For further confirming the capability of the GNP-based assay, the system was also studied by confocal laser scanning microscopy（CLSM） and classic flow cytometry（FCM） assay, and satisfactory results were obtained.

Engineering D-allulose 3-epimerase from Clostridium cellulolyticum for improved thermostability using directed evolution facilitated by a nonenzymatic colorimetric screening assay

D-Allulose 3-epimerase(DAEase)is a biocatalyst of concern for D-allulose enzymatic synthesis;however,it exhibits comparatively low thermal stability.To facilitate the directed evolution of DAEase,a nonenzymatic colorimetric assay was developed for high-throughput screening of enhanced DAEase mutants.One desirable mutant,MT4(S38F/F42N/A70P/T119P),was screened based on purifying selection of the randomly muta-genesis library and positive screening of the randomly shuffled library.Compared to the wild-type,the combi-national mutant MT4 had higher catalytic activity(1.23-fold)and showed 5,6.2,and 6.92℃ increases in T_(opt),T_(50),and T_(m) values,respectively.Furthermore,at 60℃,the t_(1/2) value of mutant MT4 was 7.30nullh,exhibiting an 11.4-fold increase in comparison to the wild-type.When the mutant MT4 was used in D-allulose production,the yield reached 28.3%,higher than the 25.1%achieved by the wild-type.Dynamic property and structural change analysis demonstrated that the enhanced properties of mutant MT4 were largely attributed to theβ4-α4 loop rigidification and active-site tunnel entrance constriction.This work expands the industrial application value of DAEase and provides a robust method as the basis for further protein engineering to achieve better performance of this enzyme.

Gold Nanoparticle-based Colorimetric Assay for Determination of Lead(Ⅱ) in Aqueous Media

The authors presented a simple colorimetric assay for the detection of toxic heavy metal lead（Ⅱ） ion（Pb2＋） Pentapeptide, cysteine-alanine-leacine-asparagine-asparagine（CALNN）, functionalized gold nanoparticles（GNPs） were aggregated in the presence of the divalent metal ion in solution by an ion-templated chelation process, which caused an easily measurable change in the absorption spectrum of the particles. Typically, mono-dispersing GNPs exhibit an absorption band at 522 nm, corresponding to a red color solution, while aggregated GNPs have it at longer wavelengths, corresponding to a purple or blue color solution. The chelation/aggregation process is reversible via the addition of a strong metal ion chelator such as EDTA. Highly selective and sensitive detection of Pb^2＋ in aqueous solution is thus provided. A detection limit of 0.1 μmol/L of Pb^2＋ was demonstrated.

New Colorimetric Method for Lipases Activity Assay in Microbial Media

A simple, rapid and precise method has been developed for determination of lipase activity in microbial media. The method is based on using phenyl acetate as substrate for lipase and determination of liberated phenol by Folin Ciocalteu reagent. Reaction mixture containing substrate 2.4 ml of phenyl acetate 165 μM in Tris HCl buffer, 0.1 M and pH 7, with 1% (v/v) Triton X-100) and 0.1 ml lipase is incubated at 40?C during 10 minutes and the absorbance was measured at 750 nm. Linearity was observed in the concentration range 0-0.8 g/L lipase.

Development of a Label-Free Colorimetric Aptasensor with Rationally Utilized Aptamer for Rapid Detection of Okadaic Acid

Okadaic acid(OA)is a typical marine toxin with strong toxicity causing diarrheic shellfish poisoning(DSP).Aptamers show great advantages in toxin detection and attract increasing attentions in the field of food analysis.In this study,a label-free col-orimetric aptasensor was constructed for visual and rapid detection of OA in shellfish.To exploit the binding capability of the anti-OA aptamer,the inherent molecular recognition mechanism of aptamer and OA was studied,based on molecular docking,fluorescent assay,and biolayer interferometry.Consistent results showed that the stem-loop near the 3’terminal of the aptamer exhibit dominate binding capacity.Based on the revealed recognition information,the aptamer was thus rationally utilized and combined with AuNPs and cationic polymer polydiallyl dimethyl ammonium chloride(PDDA)for the development of the label-free colorimetric aptasensor,in which the 3’terminal was thoroughly exposed to OA.The aptasensor provided robust performance with a linear detection range of 100-1200 nmol L-1,a limit of detection of 41.30 nmol L-1,recovery rates of 91.6%-106.2%,as well as a high selectivity towards OA in shellfish samples.The whole detection process can be completed within 1 h.To our best knowledge,this is the first time that the anti-OA aptamer was thoroughly studied,and a label-free colorimetric aptasensor was rationally designed in this way.This study not only provides a rapid detection method for highly sensitive and specific detection of OA,but also serves as a reference for the design of efficient aptasensors in the future.

Dot immunogold filtration assay for rapid detection of anti-HAV IgM in Chinese

INTRODUCTION The hepatitis A virus specific immunoglobulin M(IgM)antibody is a specific serological marker forearly diagnosis of hepatitis A..At present,themethods used at home or abroad for detecting anti-HAV IgM are RIA,ELISA and SPHAI.The dotimmunogold combination assay that has beendeveloped since 1989 is a new technique with theproperty of simple and rapid immunologicaldetection,by using the red colloidal gold particles tolabel the antibodies as indicator,and the

Monoclonal antibody-based serological detection of potato virus M in potato plants and tubers

Potato virus M(PVM) is one of the common and economically important potato viruses in potato-growing regions worldwide. To investigate and control this viral disease, efficient and specific detection techniques are needed. In this study, PVM virions were purified from infected potato plants and used as the immunogen to produce hybridomas secreting PVM-specific monoclonal antibodies(MAbs). Four highly specific and sensitive murine MAbs, i.e., 1 E1, 2 A5, 8 A1 and 17 G8 were prepared through a conventional hybridoma technology. Using these four MAbs, we have developed an antigen-coated plate(ACP)-ELISA, a dot-ELISA and a Tissue print-ELISA for detecting PVM infection in potato plants and tubers. PVM could be detected in infected potato plant tissue crude extracts diluted at 1:10 240(w/v, g mL^(–1)) by the dot-ELISA or at 1:163 840(w/v, g mL^(–1)) by the ACP-ELISA. The Tissue print-ELISA is the quickest and easiest assay among the three established serological assays and is more suitable for onsite large-scale sample detection. Detection results of the field-collected samples showed that PVM is currently widespread in the Yunnan and the Heilongjiang provinces in China. The field sample test results of the developed serological assays were supported by the results from RT-PCR and DNA sequencing. We consider that the newly established ACP-ELISA, dot-ELISA and Tissue print-ELISA can benefit PVM detection in potato plant and tuber samples and field epidemiological studies of PVM. These assays can also facilitate the production of virus-free seed potatoes and breeding for PVM-resistant potato cultivars, leading to the successful prevention of this potato viral disease.

Inactivation of Resistant Mycobacteria mucogenicum in Water:Chlorine Resistance and Mechanism Analysis

Objective To better understand the mechanism of chlorine resistance of mycobacteria and evaluate the efficiency of various disinfection processes.Methods Inactivation experiments of one strain Mycobacteria mucogenicum,isolated from a drinking water distribution system in South China were conducted with various chlorine disinfectants.Inactivation efficiency and disinfectant residual,as well as the formation of organic chloramines,were measured during the experiments.Results This strain of M.mucogenicum showed high resistance to chlorine.The CT values of 99.9% inactivation by free chlorine,monochloramine and chlorine dioxide were detected as 29.6±1.46,170±6.16,and 10.9±1.55 min（mg/L） respectively,indicating that chlorine dioxide exhibited significantly higher efficiency than free chlorine and monochloramine.It was also found that M.mucogenicum reacted with chlorine disinfectants more slowly than S.aureus,but consumed more chlorine disinfectants during longer time of contact.Lipid analysis of the cell construction revealed that 95.7% of cell membrane lipid of M.mucogenicum was composed of saturated long chain fatty acids.Saturated fatty acids were regarded as more stable and more hydrophilic which enabled the cell membrane to prevent the diffusion of chlorine.Conclusion It was concluded that different compositions of cell membrane might endow M.mucogenicum with a higher chlorine resistance.

Fast and Accurate Identification of <i>M. tuberculosis</i>Complex Using an Immunochromatographic MPT64 Antigen Detection Test

Background: A new rapid Immunochromatographic test (ICT) kit (MPT64 TB Ag Kit) for detection of MPT64 Antigen in M. tuberculosis (MTB) isolates used for rapid identification of MTB isolates developed by SD (Standard Diagnostics) Bio line, South Korea was evaluated. The ICT is a rapid, reliable and cheaper method that can be used instead of conventional biochemical tests for confirming MTB in culture isolates in resource limited laboratories. The study also evaluated the ability of ICT to detect MPT64-Antigen before the micro MGIT could signal positive. Material/Methods: A total of 450 sputum samples of individual patients were used for the study. 152 isolates of Mycobacteria were recovered from solid and liquid media. These strains were tested for the detection of MPT64-antigen. H37Rv strain was served as the positive reference control and also used for early detection of Antigen experiment. Findings: The development of bands on both test and sample region when H37Rv strain was tested were seen (MPT64 antigen positive). When 138 MTB isolates were tested, it showed a similar banding pattern indicating 100% sensitivity. MPT64 band formation was not detected in any of the 14 isolates indicating 100% specificity. Both PPV & NPV were 100%. All the isolates negative for MPT64 Ag were confirmed as MOTT by conventional bio-chemical PNBA. The H37Rv strain showed a faint band from the 2nd day onwards from inoculation till 3rd day in the earlier Antigen detection experiment. Conclusion: Rapid identification of MTB culture isolate is a pressing need for diagnosis and proceeding to perform drug susceptibility testing. MPT64 TB Ag detection ICT kit is a rapid, reliable method, good substitute for molecular identification methods, and conventional biochemical test which is time-consuming and technically demanding. The early detection of Antigen can be used as an effective tool in diagnosis.

Target-induced mimic enzyme deactivation based on mixed-node metal-organic frameworks for colorimetric assay of hydrogen sulfide

Accurate detection of hydrogen sulfide(H_(2)S)is of great significance for environmental monitoring and protection.We propose a colorimetric method for the detection of H_(2)S by the use of mixed-node Cu-Fe metal organic frameworks(Cu-Fe MOFs)as highly efficient mimic enzymes for target-induced deactivation.The Cu-Fe MOFs were synthesized by a simple solvothermal method and could catalyze the H_(2)O_(2)mediated oxidation of 3,30,5,50-tetramethylbenzidine(TMB)to oxTMB with a blue color.The presence of dissolved H_(2)S would deactivate the mimic enzymes,and then the blue color disappeared.The mechanism of the sensor was discussed by steady-state kinetic analysis.The designed assay was highly sensitive for H_(2)S detection with a linear range of 0à80 mmol/L and a detection limit of 1.6 mmol/L.Moreover,some potential substances in the water samples had no interference.This method with the advantages of low cost,high sensitivity,selectivity,and visual readout with the naked eye was successfully applied to the determination of H_(2)S in industrial wastewater samples.

Colorimetric Assay to Determine In Vitro Antibacterial Activity Against Clinical Isolates： Enhanced Activity in Damaged Chinese Masson Pine Needles

A colorimetric assay for antibacterial susceptibility testing of clinical isolates （Escherichia coli, Pseudomonas aeruginosa, Shigella dysenteriae, Staphylococcus aureus, Bacillus cereus, and Streptococcus pneumoniae） is described based on the reduction of a novel tetrazolium salt, 3-（4,5-dimethylthiazol-2-yl）-5-（3-carboxymethoxyphenyl）-2-（4-sulfophenyl）-2Htetrazolium （MTS）, in the presence of phenazine methosulfate （PMS） as an electron-coupling agent. The combination of 200 μg/mL MTS with 25 μmol/L PMS resulted in production of large amounts of formazan within 1 h of exposure. In this setting, fractions extracted from Chinese Masson pine （Pinus massoniana Lamb.） needles damaged by the pine caterpillar Dendrolimus punctatus Walker were found to have enhanced levels of antibacterial activity. These fractions, which were designated ＂Master＂, ＂Technique＂, and ＂Strength＂, were isolated and identified by reverse-phase C18 cartridge concentration, gel filtration, and affinity chromatography. Two fractions purified from healthy and undamaged needles were designated H1 and H2, respectively. For all test bacteria species. Technique produced the lowest minimal inhibitory concentrations （MICs）, ranging from 2 to 32 μg/mL, and H2 produced the highest values, with four of the six M ICs being higher than 128 μg/mL. We found that the Rmax model fitted the data well in that the r^2 ranged between 0.87 and 0.96 （median, 0.92） and no statistically significant deviations from the model were found （P= 0.23）. The median coefficient of variation of the log RC50 values and the slope m of the fitted model for all six strains among the replicates were 38 and 41%, respectively. In the course of the investigation, the physiological and functional factors involved in pest damage to plants were also explored. In summary, the MTS-PMS colorimetric assay has advantages over existing methods for the examination of antibacterial activity, and could be developed further such that it would be suitable for screening new antibiotic molecules.

Development of a colorimetric assay for rapid quantitative measurement of clavulanic acid in microbial samples

We developed a colorimetric assay to quantify clavulanic acid （CA） in culture broth of Streptomyces clavuligerus, to facilitate screening of a large number of S. clavuligerus mutants. The assay is based on a β-1actamase-catalyzed reaction, in which the yellow substrate nitrocefin （λmax=390 nm） is converted to a red product （λmax=486 nm）. Since CA can irreversibly inhibit β-1actamase activity, the level of CA in a sample can be measured as a function of the A390]A486 ratio in the assay mixture. The sensitivity and detection window of the assay were determined to be 50 μg L -1 and 50 μg L to 10 mg L-1, respectively. The reliability of the assay was confirmed by comparing assay results with those obtained by HPLC. The assay was used to screen a pool of 65 S. clavuligerus mutants and was reliable for identifying CA over-producing mutants. Therefore, the assay saves time and labor in large-scale mutant screening and evaluation tasks. The detection window and the reliability of this assay are markedly better than those of previously reported CA assays. This assay method is suitable for high throughput screening of microbial samples and allows direct visual observation of CA levels on agar plates.

The feasibility of antitumor drugs chemosensitivity testing by flow cytometry

Objective: To investigate the feasibility of chemosensitivity testing of antitumor drugs by flow cytometry in clinical applications so as to provide experimental and theoretical basis for the establishment of a novel antitumor drugs sensitivity testing and the screening of particular antitumor drugs. Methods: Detect the apoptosis rate of 12 cases of Molt-4 cell line, 57 cases of fresh clinical gastrointestinal tumor cells by Sub-G1 and Annexin V assay of flow cytometry under the effects of antitumor drugs at different times and the outcomes were compared with the ones of the MTT (3-(4,5-dimethylthiazolyl-2) -2,5-diphenyltetrazolium bromide) assay. Results: The lethality of drugs on Molt-4 cell, clinical gastrointestinal tumor cells had a positive correlation with the acting time of antidrugs by employing Annexin V, Sub-G1 and MTT assay. Drug-incurring maximum lethality of Annexin V assay was higher than MTT colorimetric assay, that of Sub-G1 was lower than MTT assay, the virtual times of Annexin V and Sub-G1 assay were obviously earlier than that of MTT colorimetric assay. Conclusion: Annexin V and Sub-G1 assay of flow cytometry can be taken as potent protocols testing anti-tumor drug chemosensitivity. Annexin V assay is featured by more sensitive, concise, reliable compared with the classical chemosensitivity testing assay of MTT colorimetric assay and it possesses clinical applied value.

A quantitative colorimetric assay of H_2O_2 and glucose using silver nanoparticles induced by H_2O_2 and UV

A simple spectrophotometric assay of H2O2 and glucose using Ag nanoparticles has been carried out. Relying on the synergistic effect of H2O2 reduction and ultraviolet （UV） irradiation, Ag nanoparticles with enhanced absorption signals were synthesized. H2O2 served as a reducing agent in the Ag nanoparticles formation in which Ag＋ was reduced to Ago by O2- generated via the decomposition of H2O2 in alkaline media. On the other hand, photoreduction of Ag＋ to Ago under UV irradiations also contributed to the nanoparticles formation. The synthesized nanoparticles were characterized by TEM, XPS, and XRD. The proposed method could determine H2O2 with concentrations ranging from 5.0× 10^-7 to 6.0× 10^-5 tool/ L The detection limit was estimated to be 2.0 × 10^-7 mol/L. Since the conversion of glucose to gluconic acid catalyzed by glucose oxidase was companied with the formation of H2O2, the sensing protocol has been successfully utilized for the determination of glucose in human blood samples. The results were in good agreement with those determined by a local hospital. This colorimetric sensor thus holds great promises in clinical applications.

Verification and Improvement of the MTT Method for in vitro HSS Bioactivity Activity Determination

To optimize the experimental conditions of MTT colorimetric assay for HSS bioactivity in vitro,we studied the optimal combination of the major conditions of the MTT assay by orthogonal test and other experiments,and compared HSS bioactivity in vitro measured by the improved MTT protocol and published MTT assay at serial protein doses.Results showed that the absorbance value(A value)of the MTT assay directly correlated with the number of human hepatoma cell lines SMMC7721.The result of orthogonal test was the number of 5×104 SMMC7721 cells/ml,culture period 6 h before adding HSS,concentration of HSS 100μg/ml,incubation time with HSS 36 h.Additionally,several experiments demonstrated the optimal combination of other conditions was 50μg MTT,incubation time for MTT 6 h,DMSO was used to dissolve the MTT formazan crystals and measured with ELISA scanner at 570 nm.The result of determining HSS bio-activity in vitro by optimized MTT protocol showed that sHSS bio-activity increased with the growth of protein dose,but decreased when it beyond a certain dose.The optimized MTT protocol was a sensitive,convenient and stable quantitative method to evaluate HSS bio-activity.

Polydopamine-Based Colorimetric Superwettable Biosensor for Highly Sensitive Detection of Hydrogen Peroxide and Glucose

Superwettable surface has broad application prospects in fabricating biosensors due to its significant enrichment effect.Here,we report a polydopamine-based colorimetric superwettable sensor that integrates superhydrophobic-superhydrophilic micropatterns for the determination of hydrogen peroxide(H_(2)O_(2))and glucose.Dopamine can be oxidized into polydopamine with the addition of horseradish peroxidase(HRP)and H_(2)O_(2),leading to the deposited spots color change from colorless to black.The concentration of target can be determined by analyzing RGB value using a smartphone software.The superhydrophobic area on the superwettable surface helps capture droplets by confining them to superhydrophilic microwells.After droplet evaporation,the analytes are concentrated in the small superhydrophilic domain,thus greatly enhancing the sensitivity.The experimental results manifested that superwettable sensor is able to detect H_(2)O_(2)with a broad linear range of 0.25μmol/L-25 mmol/L and a low limit of detection(LOD)of 0.25μmol/L by naked eye.For glucose detection,the linear range of the sensor is from 2μmol/L to 20 mmol/L and LOD is 0.69μmol/L.The superwettable sensor has been successfully applied in practical samples,including cancerous cells,milk,urine,and human serum samples with acceptable results.This superwettable sensor has several merits,such as high sensitivity,rapid response,and low sample volume in a single microdroplet,and shows great potential in manufacturing portable devices for complex biosensing applications.