Speaker
Description
About 1% of the world population is affected by an allergy for peanuts which is the most common cause of fatal-food-related anaphylaxis. Due to the high risk of exposure and the fact that doses of a few milligrams can cause such an allergic reaction, a lot of research has been done towards the detection of these immunogenic proteins. The protein Ara h1 was identified as the culprit in 95% of all allergic reactions to peanuts. To date the assays to detect the Ara h1 allergen rely on the ELISA assay, lateral flow assays and mass spectroscopy all of which are neither cheap nor fast. Aptamers, which are single DNA or RNA oligonucleotides, offer a cheaper and more stable receptor element. Amino (NH2)-terminated Ara h 1 aptamers were covalently attached to carboxylated gold surfaces and to nanocrystalline diamond with hydrogen termination using the chemical EDC coupling route. Subsequently, the functionalized surfaces were used in a setup that combines both electrochemical impedance spectroscopy and heat transfer measurements. The sensor surfaces were placed onto a copper lid which serves as a heat provider. Such an assembly was mounted onto a transparent Perspex flow cell with an inner volume of 110 ml, sealed with a miniature O-ring and fixed with screws. Two miniature thermocouples were placed at the copper backside and at 1.7 mm above the surface of the substrate in order to monitor the temperatures of the copper, T1, and of the fluid, T2, respectively. The heat flow was generated with a power resistor with a fixed resistance. For impedance spectroscopy measurements, gold electrodes served as counter electrode within a range of 100 Hz to 100 kHz. Liquids can be exchanged with a syringe-driven flow system. In this work, the aptamer functionalized surfaces were exposed to increasing amounts of Ara h1 in order to acquire response curves. Non-specific binding of the various contaminants, that a sample originating from food might bring along, are avoided by orienting the sensor face down to avoid sediments on the sensor while BSA washing is used to block any non-specific binding sites. Electrochemical impedance spectroscopy and the heat transfer method can both be used to detect Ara h1 concentrations in liquids downward to a detection limit of 3 nM. This allows the detection of Ara h1 in a dilution of 50 mg peanut butter in a 20.000 times larger volume of buffer solution.
References
[1] M. Peeters et al. Journal of Biosensors and Bioelectronics 5 (3), 155 (2014). [2] M. Peeters et al,. ACS AMI Mater. Interf. 2015, 7: 16-23.
Acknowledgements FWO project “Diamond-based impedimetric and nanophotonic biosensors for the detetection of proteins” (G.0997.11N)
