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Enzyme immunoassays are currently the methods of choice for gluten control in foods labelled as gluten free, providing a mechanism for assessing food safety for consumption by coeliac and other allergic patients. However, their limitations, many of them associated to the reactivity of the different antibodies used and their degree of specificity, have prevented the establishment of a standardised method of analysis. We explore new methods for quantitatively determining gluten content in foods based on the use of two recently described aptamers, raised against a 33-mer peptide recognised as the immunodominant fragment from α2-gliadin. The assays use the target peptide immobilised onto streptavidin-coated magnetic beads in combination with a limited amount of biotin-aptamer in a competitive format, followed by streptavidin-peroxidase labelling of the aptamer that remains bound to the magnetic beads. The enzyme activity onto the beads, measured by chronoamperometry in disposable screen-printed electrodes, is inversely related to the target concentration in the test solution. We find that while the assay using the aptamer with the highest affinity towards the target (Gli 4) achieves low detection limits (~0.5 ppm) and excellent analytical performance, when challenged in samples containing the intact protein, gliadin, it fails in detecting the peptide in solution. This problem is circumvented by employing another aptamer (Gli 1), the most abundant one in the SELEX pool, as a receptor. The proposed assays allow the convenient detection of the allergen in different kinds of food samples, including heat-treated and hydrolysed ones. The obtained results correlate with those of commercially available antibody-based assays, providing an alternative for ensuring the safety and quality of nominally gluten-free foods. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Abstract: The design and performance of an inhibition assay for the electrochemical detection of tobramycin is reported. This platform uses a monovalent system for introducing the enzyme conjugate on a tagged-aptamer that specifically recognizes the aminoglycoside antibiotic tobramycin. Compared with multivalent systems such as biotin–streptavidin, the sensitivity is greatly improved reducing the limit of detection from 5μM to 0.1μM in aqueous solution. This labeling strategy along with the use of a nuclease-resistant RNA aptamer allowed the determination of tobramycin in pre-treated human serum samples within the therapeutic range, using magnetic microparticles as a solid support for the aptassay, and differential pulse voltammetry as detection technique. [Copyright &y& Elsevier]