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Botulinum neurotoxins (BoNTs) produced by soil bacterium Clostridium botulinum are cause of botulism and listed as biohazard agents, thus rapid screening assays are needed for taking the correct countermeasures in a timely fashion. The gold standard method relies on the mouse lethality assay with a lengthy analysis time, i.e., 2–5 days, hindering the prompt management of food safety and medical diagnosis. Herein, we propose the first paper-based antibody-free sensor for reliable and rapid detection of BoNT/A and BoNT/C, exploiting their cleavage capability toward a synthetic peptide able to mimic the natural substrate SNAP-25. The peptide is labelled with the electroactive molecule methylene blue and immobilized on the paper-based electrode modified with gold nanoparticles. Because BoNT/A and BoNT/C can cleave the peptide with the removal of methylene blue from electrode surface, the presence of these neurotoxins in the sample leads to a signal decrease proportional to BoNT amount. The biosensor developed with the selected peptide and combined with smartphone assisted potentiostat is able to detect both BoNT/A and BoNT/C with a linearity up to 1 nM and a detection limit equal to 10 pM. The applicability of this biosensor was evaluated with spiked samples of orange juice, obtaining recovery values equal to 104 ± 6% and 98 ± 9% for 1 nM and 0.5 nM of BoNT/A, respectively.

In the overall scenario of precision medicine, we propose a novel paper-based lab-on-a-chip to deliver a cost-effective and easy to use sensing tool for customized administration of drugs in Alzheimer’s disease. Among several drugs, we designed the device for evaluating the efficacy of compounds (e.g. physostigmine, rivastigmine, donepezil) which are able to inhibit in a reversible way the cholinesterase enzyme. Because cholinesterase activity is peculiar to each patient, the administration of customized amount of the drug can improve the treatment efficacy and the quality of patient life, avoiding side effects due to the overdosage. In detail, we exploited Vivid™ Plasma Separation membrane to threat the whole blood sample, filter paper to load the reagents needed for the measurement, and office paper to print electrodes able to measure the butyrylcholinesterase activity, delivering a reagent free analytical tool. The calibration curve of butyrylcholinesterase obtained in blood sample provided linearity between 2 and 12 U/mL, with sensitivity of 0.050 ± 0.004 μA mL/U. The physostigmine, rivastigmine, and donepezil inhibition activities toward the butyrylcholinesterase enzyme were also measured in blood sample with linearity up to respectively 0.5 μM, 25 μM, 30 μM, and detection limits of 0.009 μM, 0.4 μM, 0.3 μM. These results demonstrate the capability of paper-based origami sensors as point of care devices to customize the drug administration in Alzheimer’s disease.

Pancreatic ductal adenocarcinoma is the predominant neoplastic disease of the pancreas and it represents the fourth most frequent cause of death in cancer-related disease, with only 8% of survivors after 5-year to the diagnosis. The main issues of this type of cancer rely on fast progress (i.e. 14 months from T1 to a T4 stage), nonspecific symptoms with delay in diagnosis, and the absence of effective screening strategies. To address the lack of early diagnosis, we report a cost-effective paper-based biosensor for the detection of miRNA-492, which is recognised as a biomarker for pancreatic ductal adenocarcinoma. To design a miniaturised, sensitive, and robust paper-based platform, an electrochemical sensor was screen-printed on office paper previously wax-patterned via wax-printing technique. The paper-based sensor was then engineered with a novel and highly specific peptide nucleic acid (PNA) as the recognition element. The formation of PNA/miRNA-492 adduct was evaluated by monitoring the interaction between the positively charged ruthenium (III) hexamine with uncharged PNA and/or negatively charged PNA/miRNA-492 duplex by differential pulse voltammetry. The paper-based biosensor provided a linear range up to 100 nM, with a LOD of 6 nM. Excellent selectivity towards one- and two-base mismatches (1MM, 2MM) or scrambled (SCR) sequences was highlighted and the applicability for biomedical analyses was demonstrated, measuring miRNA-492 in undiluted serum samples.

Altres ajuts: the ICN2 is funded by the CERCA Programme/Generalitat de Catalunya. The development of paper-based electroanalytical strips as powerful diagnostic tools has gained a lot of attention within the sensor community. In particular, the detection of nucleic acids in complex matrices represents a trending topic, especially when focused toward the development of emerging technologies, such as liquid biopsy. DNA-based biosensors have been largely applied in this direction, and currently, there are two main approaches based on target/probe hybridization reported in the literature, namely Signal ON and Signal OFF. In this technical note, the two approaches are evaluated in combination with paper-based electrodes, using a single strand DNA relative to H1047R (A3140G) missense mutation in exon 20 in breast cancer as the model target. A detailed comparison among the analytical performances, detection protocol, and cost associated with the two systems is provided, highlighting the advantages and drawbacks depending on the application. The present work is aimed to a wide audience, particularly for those in the field of point-of-care, and it is intended to provide the know-how to manage with the design and development stages, and to optimize the platform for the sensing of nucleic acids using a paper-based detection method.

In the last decades, there is a growing search for analytical strategies to ensure clinical analysis without the need of laboratory set-up and skilled personnel. Indeed, user-friendly and low-cost devices are highly valued in the era of sustainability for their capability to be applied in low-resource contexts, such as developing countries. To address this issue, herein we report a 96-well paper-based and laboratory setup-free optical platform for the detection of butyrylcholinesterase enzyme (BChE) activity in human serum. We used chromatographic paper to realize a novel analytical tool exploiting its porous structure for reagentless synthesize Prussian Blue Nanoparticles (the sensing element), as well to load all the reagents required for the measurement. The principle of BChE activity detection relies on the reaction between the enzymatic product thiocholine and Prussian Blue, giving the Prussian White with subsequently Prussian Blue's fading, detected by a common office scanner supported by ImageJ software. Using this novel paper-based optical platform, BChE activity was linearly detected in the 2–15 U/mL range with a detection limit down to 0.8 U/mL. The accuracy was successfully demonstrated by recovery study with spiked serum and by comparing the data with the gold standard method.

Herein, we propose the first three-dimensional origami paper-based device for the detection of several classes of pesticides by combining different enzyme-inhibition biosensors. This device was developed by integrating two different office paper-based screen-printed electrodes and multiple filter paper-based pads to load enzymes and enzymatic substrates. The versatile analysis of different pesticides was carried by folding and unfolding the filter paper-based structure, without any addition of reagents and any sample treatment (i.e. dilution, filtration, pH adjustment). The paper-based platform was employed to detect paraoxon, 2,4-dichlorophenoxyacetic acid, and atrazine by exploiting the capability of these different types of pesticides (i.e. organophosphorus insecticides, phenoxy-acid herbicides, and triazine herbicide) to inhibit butyrylcholinesterase, alkaline phosphatase, and tyrosinase, respectively. The degree of inhibition correlating to the quantity of pesticides was evaluated by chronoamperometrically monitoring the enzymatic activity in the absence and in the presence of pesticides by using a portable potentiostat. To improve the sensitivity, the paper-based electrodes were modified with carbon black alone in the case of platforms for 2,4-dichlorophenoxyacetic acid and atrazine detection, or decorated with Prussian blue nanoparticles for the detection of paraoxon. The paper-based device was applied for the detection of paraoxon, 2,4-dichlorophenoxyacetic acid, and atrazine at ppb level in both standard solutions and river water sample. The accuracy of this origami multiple paper-based electrochemical biosensor was evaluated in river water by recovery studies, obtaining satisfactory values (e.g. for paraoxon 90 ± 1% and 88 ± 2%, for 10 and 20 ppb, respectively). The proposed three-dimensional origami paper device allows for rapid, cost-effective and accurate pesticide detection in surface water as a result of combining filter and office papers, screen-printing, wax-printing and nanomaterial technology.

From the first investigations on peptide nucleic acids (PNA), peculiar features have been described as intrinsic of the singular chemical nature of these biomolecules. Relevant applications in the development of biosensors occurred in the last decades, exploiting them as bioreceptors. Their outstanding chemical and thermal stability as well as highly sensitive towards both complementary and non-complementary sequences, promoted them in the Olympus of biocomponents for applications in diagnostics. The intent of this review (85 ref) is to describe the last trends on the astonishing PNA technology in the biosensing field, in the view of the positive contamination with vanguard technologies including rational design and nanotechnology. In particular, electrochemical and optical biosensors exploiting PNA were reported from the literature, as well as nanostructured and paper-based biosensors for the design of microfluidics and lab-on-chip tools. (C) 2020 Elsevier B.V. All rights reserved.