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The growth of (bio)sensors in analytical chemistry is mainly attributable to the development of affordable, effective, portable, and user-friendly analytical tools. In the field of sensors, paper-based devices are gaining a relevant position for their outstanding features including foldability, ease of use, and instrument-free microfluidics. Herein, a multifarious use of filter paper to detect copper ions in bodily fluids is reported by exploiting this eco-friendly material to (i) synthesize AuNPs without the use of reductants and/or external stimuli, (ii) print the electrodes, (iii) load the reagents for the assay, (iv) filter the gross impurities, and (v) preconcentrate the target analyte. Copper ions were detected down to 3 ppb with a linearity up to 400 ppb in standard solutions. The applicability in biological matrices, namely, sweat and serum, was demonstrated by recovery studies and by analyzing these biofluids with the paper-based platform and the reference method (atomic absorption spectroscopy), demonstrating satisfactory accuracy of the novel eco-designed analytical tool.

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.

The detection of double stranded DNA (dsDNA) is often associated with the use of laboratory-bound approaches and/or with the prior generation of single stranded DNA (ssDNA), making these methods not suitable for in situ monitoring, i.e., point-of-care diagnostics. Screen-printed technology, coupled to the use of triplex forming oligonucleotides (TFO) as the recognizing probes, offers a great possibility toward the development of portable analytical tools. Moreover, the continuous demand for sustainable processes and waste lowering have highlighted the role of paper-based substrates for manufacturing easy-to-use, low-cost, and sustainable electrochemical devices. In this work, filter paper and copy paper have been utilized to produce E-DNA strips. Gold nanoparticles (AuNPs) have been exploited to immobilize the methylene blue (MB)-tagged TFO and to enhance the charge transfer kinetics at the electrode surface. Both paper-based substrates have been electrochemically characterized, and in addition, the effect of the amount of waxed layers has been evaluated. The paper-based E-DNA strips have been challenged toward the detection of three model targets, obtaining 3 and 7 nM as the detection limit, respectively, for single and double stranded sequences. The repeatability of the manufacturing (homemade) process has been evaluated with a relative standard deviation of approximately 10%. The effectiveness of the filter paper-based platform has been also evaluated in undiluted serum obtaining a similar value of the detection limit (compared to the measurements carried out in buffer solution). In addition, a synthetic PCR amplified dsDNA sequence, related to HIV, has been detected in serum samples.

We report a reagentless, electrochemical sensor for the detection of double-stranded DNA targets that employs triplex-forming oligonucleotides (TFOs) as its recognition element. These sensors are based on redox-tagged TFO probes strongly chemisorbed onto an interrogating gold electrode. Upon the addition of the relevant double-stranded DNA target, the probe forms a rigid triplex structure via reverse Hoogsteen base pairing in the major groove. The formation of the triplex impedes contact between the probe's redox moiety and the interrogating electrode, thus signaling the presence of the target. We first demonstrated the proof of principle of this approach by using a well-characterized 22-base polypurine TFO sequence that readily detects a synthetic, double-stranded DNA target. We then confirmed the generalizability of our platform with a second probe, a 19-base polypyrimidine TFO sequence that targets a polypurine tract (PPT) sequence conserved in all HIV-1 strains. Both sensors rapidly and specifically detect their double-stranded DNA targets at concentrations as low as ~10 nM and are selective enough to be employed directly in complex sample matrices such as blood serum. Moreover, to demonstrate real-world applicability of this new sensor platform, we have successfully detected unpurified, double-stranded PCR amplicons containing the relevant conserved HIV-1 sequence.

A new method for aflatoxin B (AFB) determination is proposed. The AFB determination is based on acetylcholinesterase (AChE) inhibition, and the AChE residual activity is determined using the colorimetric method (Ellman's method). Cholinesterases (ChEs) from various sources were tested using AFB1 as reference aflatoxin. AChE from electric eel has shown the highest sensitivity to AFB1, and it was chosen for the rest of the work. To select and optimize the analytical procedures, an investigation on the type of AChE inhibition by AFB1 was carried out. The AChE degree of inhibition by AFB1 was independent of the incubation time and the enzyme concentrations, showing the reversibility of the inhibition. This reversibility of the inhibition permits a rapid analysis of AFB1, requiring only 3 min. For the development of the AFB1 assay, the pH, the time of reaction, temperature, and substrate concentration were evaluated and optimized. The linear range of 10-60 ng mL-1 was determined. To evaluate the selectivity of this method, the cross-reactivity with other aflatoxins such as aflatoxin B2, aflatoxin G1, aflatoxin G2, and aflatoxin M1 was investigated. Finally, the suitability of the assay for AFB1 quantification in barley was evaluated. This study shows a new approach to detect aflatoxins based on enzyme inhibition and has advantages such as the ease of use, rapidity, and cost effectiveness. Thus, it could find a possible use as a screening method for this type of mycotoxins.