A novel AIEgen-Based probe Integrated with split aptamer Conjugates for adenosine triphosphate detection in beer.

Aptamer-based strip assay is an easy, highly efficient and low-cost detection method, which has been developed and easily applied to onsite detection. A new sensitive sandwich dipstick assay for adenosine triphosphate (ATP) detection was successfully developed based on specific recognition between split aptamer fragments and the target. In this method, the thiolated aptamer was first conjugated to the surface of gold nanoparticles (AuNPs), while the biotin-aptamer was immobilized on the surface of a nitrocellulose filter in the test line. In the presence of ATP, the thiol-aptamer/ATP/biotin-aptamer complexes were generated, which led to an obvious increase in optical signals at the test line. Under the optimal determination conditions, an excellent linear logarithmic response to the ATP concentration was obtained within the range of 0.5μM to 5mM. The limit of detection (LOD) of 0.5μM was reached at a signal-to-noise ratio of 3. The dipstick assay showed a good average recovery of 96-108% with the RSD of less than 20% in urine samples. The proposed method exhibited high specificity against other nucleotides such as the uridine triphosphate (UTP), cytidine triphosphate (CTP), and guanosine triphosphate (GTP). The results indicated that the dipstick strip may be considered as an inexpensive screening tool for onsite ATP determination. Graphical Abstract A simple split aptamer fragments based sandwich-type dipstick assay was developed for ATP detection.

Adenosine triphosphate (ATP) is crucial for cellular activity. The need for ATP detection in the field of biomedicine is rapidly increasing. Several biosensor-based approaches have been developed as a result of the growing demand for ATP detection. An electrolyte-insulator-semiconductor (EIS) sensor is a type of field-effect device that has the ability to detect surface-potential changes with a specific level of sensitivity. In this study, a label-free ATP detection biosensor based on poly(amidoamine)-modified EIS sensors was developed, in which an ATP-sensitive aptamer (Apt) was utilized as the sensitive element and the EIS sensor was used as the transducer. It is possible to monitor the binding of charged molecules, such as aptamers using EIS sensors in a label-free manner with a straightforward setup. To improve the coupling efficiency of Apt with the EIS sensor, a positively charged polyelectrolyte, i.e., poly (amidoamine) (PAMAM) dendrimers, was utilized to modify the surface of the EIS sensors to attach the negatively charged Apt through electrostatic attraction. The adsorptive binding of Apt and ATP results in a change in the capacitance of the EIS sensor. During the process of surface modification, the electrochemical measurements of capacitance-voltage (C-V) curves and constant-capacitance (ConCap) characteristics were utilized as indicators for the corresponding processes of EIS sensor surface modification. The measurement results indicated that this biosensor was able to detect ATP with high sensitivity and good specificity. The detection range of ATP was from 0.1 nM to 100 nM and the detection limit was as low as 0.12 nM. This biosensor has the potential to be utilized in the detection of ATP in the surrounding microenvironment of cells and tissues, with promising prospects for application in the field of biomedicine such as energy and metabolism monitoring of cells and tissues.

Aptamers, which are in vitro selected functional oligonucleotides, have been employed to design novel biosensors (i.e., aptasensors) due to their inherent selectivity, affinity, and their multifarious advantages over traditional recognition elements. In this work, we reported a multifunctional reusable label-free electrochemical biosensor based on an integrated aptamer for parallel detection of adenosine triphosphate (ATP) and alpha-thrombin, by using electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). A Au electrode as the sensing surface was modified with a part DNA duplex which contained a 5'-thiolated partly complementary strand (PCS) and a mixed aptamer (MBA). The unimolecular MBA contained small-molecule ATP binding aptamer (ABA) and also protein alpha-thrombin binding aptamer (TBA). Thus, the aptasensor could be used for detection of ATP and alpha-thrombin both. The detection limit of ATP was 1 x 10(-8) M, and its detection range could extend up to 10(-4) M, whereas the detection limit of alpha-thrombin was 1 x 10(-11) M, and its detection range was from 1 x 10(-11) to 1 x 10(-7) M. Meanwhile, after detecting alpha-thrombin, the sensing interface could be used for ATP recognition as well. The aptasensor regeneration could be realized by rehybridizing of the MBA strand with the partly complementary strand immobilized on the Au surface after ATP detection or by treating with a large amount of ATP and then rehybridizing the MBA strand with the partly complementary strand immobilized on the Au surface after alpha-thrombin detection. The aptasensor fabricated exhibited several advantages such as label-free detection, high sensitivity, regeneration, and multifunctional recognition. It also showed the detectability in biological fluid. Therein it held promising potential for integration of the sensing ability such as the simultaneous detection for multianalysis in the future.

Use of a small molecule as an initiator for interchain staudinger reaction: A new ATP sensing platform using product fluorescence

The cationic water-soluble poly(diethynylfluorene-co-diethynylcarbazole-co-diethynylbenzothiadiazole) (P1–P3) were synthesized and used for adenosine triphosphate (ATP) detection. The incorporation of diethynylbenzothiadiazole (DBT) into the polymer backbone offers dual emissive polyelectrolytes with both blue and red emission bands in the dilute solution resulting from incomplete intramolecular energy transfer. In the solution of P2, addition of ATP can quench the emission of fluorene-co-carbazole units of P2 more efficiently than that of DBT segments, which causes the fluorescent color change from blue to red. The asymmetric quenching of the blue and red emission bands in the presence of ATP might become a possible method for a real-time ATP detection. In addition, when ATP is cleaved into fragments by enzyme alkaline phosphatase (ALP), the blue emission of P2 can be recovered. Thus, it is possible to assay the enzyme activity by triggering the change in color and intensity of the emission of the solution of P2.

Adenosine triphosphate (ATP) is a critical intracellular energy currency that plays a key role in various cellular processes and is closely associated with numerous diseases. Similarly, biothiols such as glutathione (GSH), cysteine (Cys), and homocysteine (Hcy) are integral to many physiological and pathological processes due to their strong redox properties. Simultaneous discrimination and detection of ATP and biothiols offer valuable insights into the pathogenesis of conditions such as epilepsy and liver injury. This study introduces the first fluorescent probe, BCR, designed for multifunctional detection of ATP, GSH, Hcy, and Cys. With outstanding optical properties, excellent biocompatibility, high selectivity, and superior sensitivity, probe BCR enables effective imaging of ATP and biothiol dynamics in vivo. Moreover, probe BCR successfully visualizes changes in ATP, GSH, Hcy, and Cys levels in a PTZ‐induced epileptic zebrafish model and an APAP‐induced mouse liver injury tissue section model. These findings underscore the significant potential of probe BCR for early disease diagnosis and therapeutic applications.

A microfluidic device for adenosine triphosphate (ATP) was checking of fish freshness. Sample solutions containing ATP and the other solution containing an enzyme substrate were handled in the form of plugs formed in a network of flow channels. To simplify the structure and procedure for mixing solutions, a wider portion was formed in a flow channel to merge the plugs. A row of plugs were ejected from each of three branched flow channels, merged at the wider portion, and transported to an ATP biosensor located in the lower stream. ATP was detected using two enzymatic reactions involving glycerol kinase (GK) and glycerol-3-phosphate oxidase (G3PO). The current observed with solutions of three different ATP concentrations showed different values, demonstrating that the device can be used for multi-point sensing. Furthermore, the detection of ATP in extracts from tuna was carried out and a distinct increase in current was observed as in the case of standard solutions.

A label-free fluorescence aptasensor was studied for adenosine triphosphate (ATP) detection that combines low biosensor toxicity with a simplified preparation process. In this study, the aptamer of the gold nanoparticle@aptamer@carbon quantum dot nanostructure could specifically identify ATP, resulting in the change of the fluorescence signal. In order to analyze the performance of the sensor, the effects of the carbon quantum dot (CQD) concentration and centrifugal rate on the stability of the probe were investigated. The results show that the sensor was superior under the 220 µl CQD volume and 2000 rpm centrifugal rate. Furthermore, the linear relationship between the change of the fluorescence signal and ATP concentration is Y = 359.747 + 0.226X within the volume range of 20 µM–280 µM. The correlation coefficient is 0.98, and the detection limit is 20 µM. No obvious fluorescence change was observed in solutions containing other common ions. On the basis of no pollution and simplicity, this sensor demonstrates great potential as a low-cost diagnostic tool for the detection of various targets, particularly for use in the fields of food safety and biomedical diagnostics.

Graphene foam field-effect transistor for ultra-sensitive label-free detection of ATP

Nucleic acid-mediated nanomachines have significant potential in biomedical applications but new approaches that link molecular recognition of proteins to change in nucleic acid structure and function are required. Here, a split DNA aptamer is integrated into G-quadruplex tweezers, which close in the presence of the malaria biomarker protein Plasmodium falciparum lactate dehydrogenase (PfLDH). Closing of the tweezers enables G-quadruplex hemin mediated peroxidase activity, which can be observed colorimetrically. The PfLDH aptamer is split within an asymmetric internal loop and incorporated into the tweezers maintaining aptamer binding capability. Spacing between the G-quadruplex structure and split aptamer, together with extent of complementarity, is found to be critical for optimization to enhance catalytic performance. The integrated split aptamer is observed to maintain the high specificity to Plasmodium falciparum lactate dehydrogenase of the parent aptamer. Split aptamer approaches have significant potential to functionalize nucleic acid nanostructures for protein molecular recognition.

Antifouling aptasensor for the detection of adenosine triphosphate in biological media based on mixed self-assembled aptamer and zwitterionic peptide

Pathogens such as porcine epidemic diarrhea virus (PEDV), porcine reproductive and respiratory syndrome (PRRSV), and E. coli are known to spread by contaminated vehicles and equipment. Pork producers have adopted trailer wash policies where each trailer is washed, disinfected, and dried before it can return to a farm. Cleanliness of livestock trailers after washing is determined by visual inspection rather than any objective method. Adenosine triphosphate (ATP) bioluminescence is used in many industries to provide real-time feedback on surface cleanliness through the detection of ATP from organic sources. That same technology may provide trailer wash facilities a way of objectively characterizing a livestock trailer's suitability to return to a farm after washing. Two ATP luminometers (3M Clean-Trace and Neogen AccuPoint) were used to estimate the correlation between ATP bioluminescence readings and aerobic bacterial plate counts (APCs) from sampled surfaces and to determine locations within a livestock trailer that can accurately estimate surface cleanliness. Five locations in livestock trailers were evaluated. Those locations included the nose access door (NAD), back door flush gate, rear side access door (RSAD), belly flush gate (BFG), and belly side access door (BSAD). There was a positive log-log association between the two luminometers (r = 0.59, P < 0.01). Every log unit increase in one unit, resulted in a 0.42 log increase (P < 0.01) in the other unit. ATP can come from bacteria, yeasts, molds, and manure. There was a poor association (r ≥ 0.10, P ≥ 0.02) between APCs and the ATP luminometers. Still, an increase in relative light units (RLUs) resulted in a corresponding increase in colony-forming units. The greatest area of surface contamination measured by APC was the NAD. RLUs were also greater in the NAD compared to the RSAD, the BFG, and the BSAD (P ≤ 0.01). Because APCs and luminometer RLUs provided similar outcomes, statistical process control charts were developed to determine control limits for RLUs. This provides real-time feedback to trailer wash workers in determining cleanliness outcomes for livestock trailers. These data suggest that ATP bioluminescence can be a reliable method to monitor cleaning effectiveness in livestock trailers. Bioluminescence is a monitoring tool that should be used in conjunction with microbial methods to monitor procedures for cleaning and disinfection.

There are many reasons for testing cell viability. Simple tinctorial assays, such as trypan blue exclusion, have their place, but luminescence assays based on the detection of adenosine triphosphate (ATP) are particularly useful, since it is possible to detect the ATP present in fewer than 20 cells/mL using optimized reagents. These same reagents can also accurately determine the ATP from 2 × 107 cells/mL in an adjacent well or test tube with a completely linear relationship between cell number and light output, provided that the cells maintain a relatively invariant ATP content. Similar methods are used for bacteria and eukaryotic cells, but this chapter is restricted to the consideration of eukaryotic cells. Luminescence measurement of ATP levels uses the following reaction:KeywordsLymphocyte Transformation TestTrypan Blue SolutionAutomate PipetUniversal TubeMaximum Nontoxic ConcentrationThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

In the present study, based on multifunctional Dual-Hairpin DNA structure, a simple, fast and high sensitive assay for the detection of DNA, thrombin and adenosine triphosphate (ATP) was demonstrated. DNA sequence labeled with methylene blue (MB), which was designed as single-stranded DNA (ssDNA) matching with target DNA, thrombin, or ATP aptamer, hybridized to the adjunct probe and formed the dual-hairpin structure on the electrode. With the hybridization of adjunct probe and the hairpin-like capture probe in the stem region, the dual-hairpin was formed with outer and inner hairpins. By the conjugation of the target probe with the adjunct probe in the outer hairpin, the adjunct probe divorced from the dual-hairpin structure. The adjunct probe with signal molecules MB, attaching near or divorcing far from the electrode, produced electrochemical signal change and efficient electron transfer due to the fact that it was in proximity to the electrode. However, upon hybridization with the perfect match target, the redox label with the target probe was forced away from the modified electrode, thus resulting in the change of the Dual-Hairpin DNA conformation, which enables impedance of the efficient electron transfer of MB and, consequently, a detectable change of the electrochemical response. In addition, another highlight of this biosensor is its regenerability and stability owing to the merits of structure. Also, based on this Dual-Hairpin platform, the detection limits of DNA, thrombin, and ATP were 50 nM, 3 pM, and 30 nM, respectively. Moreover, this pattern also demonstrated excellent regenerability, reproducibility, and stability. Additionally, given to its ease-of-use, simplicity in design, easy operations, as well as regenerability and stability, the proposed approach may be applied as an excellent design prompter in the preparation of other molecular sensors.

The present study focuses on the improvement of limitations of the bioluminescent ATP assay. An attempt was made to improve on the following: non-selective detection, limited cell population density quantification, and to distinguish between bioluminescent noises data and cell ATP, as a suitable technique for coliforms and Escherichia coli detection, as well as other cell strains. Results were achieved by combining growth kinetics of the cell population exponential phase, in 2% brilliant green lactose bile broth with bioluminescent assay, based on the reaction catalyzed by firefly luciferase. The equations applied to the description of kinetics on exponential growth phase were able to estimate the relative light units (RLU0) corresponding to initial adenosine triphosphate (ATP0) concentration. The developed equations also help to distinguish between background RLU and RLU0 corresponding to cell ATP, and to analyze the kinetics with and without lag period in order to estimate RLU0. The results demonstrated a good correlation with data obtained by standard methods for coliforms detection. The quantification of E. coli and coliforms could be performed within a period of 6 h at 3.24 x 105 and 1.12 x 103 CFU/mL from the calculation of initial ATP concentrations at 19 and 0.067 pmol/mL, respectively. Monitoring of growth kinetic parameters along with the bioluminescent detection of microorganisms in the selective medium during shorter test time has potential for their selective and sensitive quantification. The mathematical approach may be extended to the analysis of experimental data obtained by other techniques based on the cell metabolite detection. Key words: Bioluminescence, coliforms, Escherichia coli, exponential phase of growth, adenosine triphosphate (ATP) detection.