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The COVID-19 pandemic highlighted lateral flow immunoassay (LFIA) strips as the most known point-of-care (POC) devices enabling rapid and easy detection of relevant biomarkers by nonspecialists. However, these diagnostic tests are usually associated with the qualitative detection of the biomarker of interest. Alternatively, electrochemical-based diagnostics, especially known for diabetes care, enable quantitative determination of biomarkers. From an analytical point perspective, the combination of the two approaches might represent a step forward for the POC world: in fact, electrochemical transduction is attractive to be integrated into LFIA strips due to its simplicity, high sensitivity, fast signal generation, and cost effectiveness. In this work, a LFIA strip has been combined with an electrochemical transduction, yielding an electrochemical LFIA (eLFIA). As a proof-of-concept method, the detection of prostate-specific antigen has been carried out by combining a printed-electrochemical strip with the traditional LFIA tests. The electrochemical detection has been based on the measurement of Au ions produced from the dissolution of the gold nanoparticles previously captured on the test line. The analytical performances obtained at LFIA and eLFIA were compared, highlighting how the use of differential pulse voltammetry allowed for a lower detection limit (2.5-fold), respectively, 0.38 and 0.15 ng/mL, but increasing the time of analysis. Although the correlation between the two architectures confirmed the satisfactory agreement of outputs, this technical note has been thought to provide the reader a fair statement with regard to the strength and drawbacks about combining the two (apparently) competitor devices in a diagnostics field, namely, LFIA and electrochemical strips.

The design, preparation and evaluation of molecularly imprinted polymers for roxarsone (4-hydroxy-3-nitrophenylarsonic acid), an organo-arsenic swine and poultry feed additive, using bi-substituted ureas and squaramide receptors as the functional monomers, are demonstrated. Pre-polymerisation studies of the template–monomer complexation performed by 1H NMR experiments show that squaramide-based monomers provide association equilibrium constant values higher than urea-based monomers. Equilibrium rebinding experiments in methanol show that two squaramide-based materials have good molecular recognition properties towards roxarsone, with high affinity (Keq= 16.85 × 103L mol−1and 14.65 × 103L mol−1, respectively), high imprinting factors (4.73 and 3.64 respectively) and good selectivity towards two roxarsone-related compounds, acetarsone (3-acetamido-4-hydroxyphenylarsonic acid) and nitarsone (4-nitrophenylarsonic acid). Polymer MIP-SQ2 was successfully used to setup an experimental protocol for the direct solid phase extraction of roxarsone from surface water samples. The method gives clean HPLC traces, with recoveries between 91% and 95% at concentration levels of 5.0, 10, and 25 mg L−1. Sample preconcentration with good recoveries between 87% and 97%, are shown, confirming that it is possible to employ the developed materials to measure roxarsone down to 1 μg L−1in water samples.

Stable and efficient conjugates between antibodies and gold nanoparticles (GNP-Ab) are sought to develop highly sensitive and robust biosensors with applications in medicine, toxicology, food safety controls, and targeted drug delivery. Several strategies have been proposed for directing the antibody attachment to GNPs thus preserving antibody activity, including covalently coupling the antibody to a polymer grafted on GNP surface and exploiting the high affinity of bioreceptors as mediators for the binding. Both approaches also allow for shielding GNPs with a protective layer that guarantees the robustness of the conjugate. Notwithstanding, antibodies freely adsorb to GNP with high binding efficiency. The nonspecific adsorption is far more simple, fast, and inexpensive than any mediated coupling. Therefore, it is preferred for most applications, although it is considered to produce GNP-Ab with a limited activity. In this work, we compared three strategies for producing GNP-Ab, such as (i) covalent coupling mediated by a chemical layer, (ii) affinity-based binding mediated by a biomolecular layer composed of Staphylococcal protein A, and (iii) direct attachment via adsorption. The so-prepared GNP-Ab were employed as probes in a colorimetric lateral flow immunoassay (LFIA) for measuring salivary cortisol as a model biosensor that relies on the use of active GNP-Ab conjugates. Unexpectedly, the biosensors fabricated using the three probes were completely comparable in terms of their ability to measure salivary cortisol. Furthermore, we observed that the sensitivity of the LFIA primarily depended on the amount of the antibody bound to GNPs rather than on the method by which it was bound. The probes prepared using both the direct adsorption approach and mediated coupling via the biochemical mediator enabled development of point-of-care devices for the fast, sensitive, and reliable measurement of human salivary cortisol.

The development of reliable molecularly imprinted sorbent assays is ensured by in-depth knowledge of the binding between the tracer, conventionally based on a template analogue conjugated to an enzyme, and the imprinted polymer used as a recognition element. To this end, the binding properties of a cortisol-3-(O-carboxymethyl)oxime-horseradish peroxidase conjugate to cortisol-imprinted microparticles previously adsorbed at the bottom of microplates were assessed. The effect of different blocking agents as well as of different percentages of Tween 20 in the working buffer were investigated in order to minimise the non-specific binding of the enzyme tracer to the adsorbed imprinted microparticles. The capability of the enzyme tracer to bind the imprinted solid phase and to compete with free cortisol was assessed by measuring the apparent equilibrium dissociation constant, KD(39.7 ± 13.5 pmol L−1), and the apparent binding site concentration, Bmax(21.7 ± 4.3 nmol L−1), whereas the IC50value for the cortisol competition curve was found to be 5.32 ± 1.15 ng mL−1. Moreover, binding selectivity measured for several cortisol-related steroids confirmed the experimental results previously published for cortisol-imprinted polymers. A competitive assay for the determination of cortisol in human saliva was developed with a limit of detection of 1.02 ng mL−1, providing quantitative results comparable to those of a commercial ELISA.

Mycotoxins are toxic compounds produced by various moulds, which can affect a variety of crops. Due to their high toxicity and wide diffusion, mycotoxins constitute a severe risk for human health, therefore maximum tolerance levels in food and feed products have been set up all over the world and analytical controls are mandatory for many commodities. Despite validated analytical methods for assessing mycotoxin contamination already existing, a number of papers describing new methods of extraction, identification or measurement appear daily in literature. Nevertheless, the extraction and determination of such contaminants in food and feed is a topic of constant and increasing interest, as also attested by the number of related patents which have been applied for in the last few years. Scientific papers dealing with mycotoxin occurrence, potential risk and determination have recently been reviewed. Nevertheless, the objective of this review is to focus on patent activity rather than on the scientific breakthroughs on this subject. Therefore, the most recent patents regarding the whole analytical protocol to measure mycotoxins in food, starting from 2006 to date are presented and discussed. The possibility of a technology transfer for the various innovations presented is also discussed as are future developments in the field.

A method for molecularly imprinted SPE of banned Sudan azo‐dyes from food samples was investigated. The molecularly imprinted polymer was obtained by suspension polymerization using 1‐(4‐chlorophenyl)azonaphthalen‐2‐ol as the mimic template. The molecular recognition properties of imprinted beads were evaluated for use as a SPE sorbent, in order to develop a selective extraction protocol for the Sudan class of dyes. The optimized extraction protocol resulted in a reliable molecularly imprinted SPE (MISPE) method suitable for HPLC analysis. It was selective for the main analyte, Sudan I, and the related azo‐dyes Sudan II, III, IV, Sudan Red B, and Sudan Red 7B, while the permitted azo‐dyes Allura Red AC, Neococcin, and Sunset Yellow FCF were not extracted. The method was tested for Sudan I, II, III, and IV in five different food samples (hot chilli pepper, hot chilli tomato sauce, sausage, tomato sauce, and hard boiled egg yolk) at three concentration levels (15, 100, and 300 μg/g). It demonstrated itself to be insensitive to the presence of different complex matrices, precise, accurate, and with good recovery rates (85–101%). The LOD and LOQ were satisfactory for most analytical determinations.

Molecular imprinted polymers are man-made materials with predetermined selectivity towards one or more analytes. They have been used as selective stationary phases in several analytical separative techniques. This review discusses the possible uses of imprinted polymers, with a particular emphasis on their use as high pressure and electrochromatographic stationary phases for resolution of optical enantiomers, materials for fast screening of libraries and selective solid-phase extraction for sample clean-up and preconcentration.

A coating procedure based on the physical adsorption of hydroxypropyl cellulose onto the wall of a capillary column has been successfully used for the separation of DNA fragments up to 500 bp. The method uses a running Tris‐phosphate‐EDTA buffer containing 2‐hydroxyethyl cellulose as sieving polymer. The separation procedure shows good reproducibility (measured as RSD%) for consecutive runs (< 0.64), for different days (< 1.15) and capillaries (< 2.15), short analysis times, and a long coating lifetime. Good reproducibility and efficiency are even achieved by performing the separation in the presence of additives such as ethidium bromide and mannitol. The method is applied to the detection of GMOs in soybean and maize meals with an accurate evaluation of the length of DNA sequences, previously amplified by polymerase chain reaction.

The different behaviour of two isozymes (IsoA and IsoB) of catechol 1,2-dioxygenase (C1,2O) from Acinetobacter radioresistenss13 on a hydrophobic interaction, Phenyl-Sepharose chromatographic column, prompted us to investigate the role of superficial hydrophobicity on structural-functional aspects for such class of enzymes. The interaction of 8-anilino-1-naphtalenesulphonate (ANS), a fluorescent probe known to bind to hydrophobic sites in proteins, revealed that the two isoenzymes have a markedly different hydrophobicity degree although a similar number of hydrophobic superficial sites were estimated (2.65 for IsoA and 2.18 for IsoB). ANS is easily displaced by adding the substrates catechol or 3-methylcatechol to the adduct, suggesting that the binding sites are in the near surroundings of the catalytic clefts. The analysis of the hydropathy profiles and the possible superficial cavities allowed to recognize the most feasible region for ANS binding. The lower hydrophobicity detected in the near surroundings of the catalytic pocket of IsoB supports its peculiarity to lose the catalytic metal ions more easily than IsoA. As previously suggested for other metalloenzymes, the presence of more hydrophilic and/or smaller residues near to the active site of IsoB is expected to increase the metal ligands mobility thus increasing the metal ion dissociation rate constants, estimated to be 0.078 h−1and 0.670 h−1for IsoA and IsoB respectively. Abbreviations:C1,2O – catechol dioxygenase; ANS – 8-anilino-1-naphthalenesulphonate; PHO – phenol hydroxylase oxygenase