Despite some recent developments on the portable on-site sensor of Aflatoxin B1 (AFB1), the complex and expensive preparation of recognition elements have still limited their wide applications. In this paper, using the fast, low-cost, and stable recognition of aptamer DNA-AFB1, a portable aptasensor was constructed for the on-site detection of AFB1 in food matrixes, with the readout of personal glucose meter (PGM) and DNA walking machine for signal probe separation. In such an assay protocol, the target could trigger the DNA walker to autonomously move on the electrode surface, propelled by unidirectional Pb-specific DNAzyme digestion, which could amplify the signal and separate the signal probe as well for further quantification by the PGM. Under optimized conditions, the increase of PGM signal was relative with the concentration of AFB1 ranging from 0.02 to 10 nM and the low limit of detection (LOD) was 10 pM (S/N = 3). With the features of portability, and cheapness, the presented user-friendly method could be extended to various other analytes for wide point-of-care applications.

In this paper, a novel colorimetric method for the detection of alkaline phosphatase (ALP) and pyrophosphate (PPi) was designed based on a Cu(2+)-horseradish peroxidase (HRP)-3,3',5,5'-tetra-methylbenzidine (TMB)-H2O2 system. In the presence of ALP, l-ascorbic acid-2-phosphate (AAP) could be hydrolyzed to ascorbic acid which could reduce Cu(2+) to Cu(+) to inhibit the enzymatic activity of HRP in the colorimetric system. The change in absorbance was found to be proportional to the ALP concentration with a linear detection range and a limit of detection of 5.4 mU mL(-1). In the presence of PPi, because Cu(2+) was chelated by PPi, the conversion of Cu(ii) by AA was effectively inhibited. The color of the HRP-TMB-H2O2 system with Cu(2+) showed blue. The HRP-TMB-H2O2 system with the Cu(2+) colorimetric system could also detect PPi with a satisfying result. In summary, this method possesses sensitivity, reproducibility, and cost-effectiveness without labelling and separation and the use of a colorimetric method is more in line with the requirements of on-site detection and green chemistry.

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Determination of nucleotide kinase activity is valuable due to its importance in regulating nucleic acid metabolism.

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Thioflavin T (ThT), as one of the most exciting fluorogenic molecules, boasts the "molecular-rotor" ability to induce DNA sequences containing guanine repeats to fold into G-quadruplex structures. It has been demonstrated to sense this change by its remarkable fluorescence enhancement. In this work, taking T4 polynucleotide kinase (PNK) as a model, the ThT/G-quadruplex based platform and λexonuclease (λexo) cleavage reaction were combined to design a label-free "turn-on" strategy for fast, simple and accurate detection of T4 PNK activity and its inhibition. In the presence of T4 PNK, the designed thioflavin T based molecular beacon (TMB) DNA probe could be phosphorylated and then digested by the cleavage of λexo, releasing the G-quartets. These then bound to ThT to form ThT/G-quadruplexes with an obvious fluorescence generation, for the "turn-on" detection of T4 PNK. In comparison to traditional methods, the proposed TMB probe is convenient, requiring no sophisticated labeling and separation processes and displaying high analytical performance. It exhibits a satisfying detection result for the activity of T4 PNK with a low detection limit of 0.001 U mL(-1). This is not only meaningful for further research on disease-related biochemical processes, but also valuable for molecular-target therapies.