Aptamer-based biosensor for food allergen determination using graphene oxide/gold nanocomposite on a paper-assisted analytical device

The detection of allergens in food are currently conducted by techniques that are time consuming and complicated which can deter consistent sampling for allergens, which could potentially cause an anaphylactic shock in the consumer by cross contamination. The need for a technique that is rapid, on site, cost effective, disposable, highly sensitive and accurate to identify these molecules urges the development of a point of care device. The aim of this work is to develop a microfluidic paper assisted analytical device (micro PAD) using hydrophobic channels, set by a wax printer on filter paper, and functionalized gold nanoparticles (AuNP) to help identify the allergens arachin (Ara h 1) for peanuts, beta lactoglobulin (beta LG) for milk, and tropomyosin (Pen a 1) for shrimp and other shellfish presence by a colorimetric test. Synthesized AuNP were conjugated with biotinylated aptamers, using the biotin streptavidin interaction, to make the specific detection of target allergens. Functionalized AuNP are incubated with the sample and are absorbed by graphene oxide (GO), creating GO AuNP complexes, if the aptamers have not become structured due to conjugation with allergenic proteins. The microPAD device is used to filter the resultant mixture which provides superior sensitivity to detect the allergens present down to the nanogram range (allergens were measured from 25 nM to 1000 nM with a LOD of 7.8 nM, 12.4 nM and 6.2 nM for peanut, milk and shrimp allergens respectively), in contrast to the microgram range of commonly used enzymatic immunoassays. The simple color indicator, varying from clear to pink in the presences of allergens allows the readout to be utilized without the need for highly specific equipment or training. Alternatively, the results can be quantified by taking a picture and measuring the color. This presented microPAD can provide results in real time and has the potential to become a rapid, low cost, and accurate portable point of care device to avoid cross-reactivity of food borne allergens.