Food Safety Diagnostics
Food safety becomes an emerging issue since bovine spongiform encephalopathy (BSE) and dioxins scandals occurred in the late 1990s. Public confidence in healthy food reached the minimum level and European policy makers spent many effort to improve the healthiness of food products and to enhance the level of consumer’s health protection. Stringent food safety and quality analysis represent the keystone of the plan of action in order to reassess consumer confidence and trust in food and food industries. It is a global issue that requires the concerted efforts of all the actors of the entire supply chain, research organism, control authorities and politicians. Food plants are heavily constrained to maintain a rigorous monitoring system for food analysis to ensure compliancy with legislation and rules as laid out by governments and regulatory authorities.
In this context, it is evident the importance of detection systems, which have to be accurate and sensitive, cheap and preferably portable for on-site testing. Biosensors represent rapid screening tools and their use as detection platform is expected to increase continually. A biosensor is a device that uses specific biochemical reactions mediated by isolated enzymes, immunosystems, tissues, organelles or whole cells to detect chemical compounds usually by electrical, thermal or optical signal.
It is composed by three main components: the biological recognition element, the transducer and the signal readout system. Once the analyte interacts with the bioreceptor, the transducer translates the recognition event in a measurable signal that is then converted into an appropriate output. According to the transduction method, biosensors are typically classified as optical, mechanical and electrochemical sensor platforms. Such innovative devices offer advantages over current analytical methods: they show good selectivity, low cost, portability, ease of use. For these reasons they represent a valid alternative to conventional method used in food industry for hazardous food contaminants detection such as mycotoxins, heavy metals, pesticides, antibiotic residues and pathogenic microorganism.
The global objective of this research activity is the development of high sensitive microcantilever biosensors for food hazard analysis. They represent a promising label-free sensing platform because they are highly sensitive, cheap, versatile, fast and ease of use.
Microcantilevers (MC) are single-side clamped microbeams that can be activated with a specific antibody in order to bind target molecules. It is possible to weigh the total mass of analyte captured by the sensor measuring the resonance frequencies before and after binding events. The research activity focused on detection of common food hazard agents like carcinogenic mycotoxins (Ochratoxin A and Aflatoxins), pathogen microorganisms (Salmonella spp. and Lysteria monocytogenes), growth promoting agent (17-β-estradiol) and allergens.
MC resonators were fabricated starting from Silicon-On-Insulator (SOI) wafers, using bulk and surface micromachining processes based on optical lithography. Each array comprises 11 MCs, with length, width and thickness respectively in the range of 460–600 μm, 50–70 μm, 5–10 μm. Resonance frequencies are monitored using Cantired, a highly automated system made by Microla Optoelectronics s.r.l.
A SEM image of a microcantilever array. The characterization set-up.
To date control strategies in detecting anabolic agents for promoting growth of food producing animals are mainly related to screening techniques based on immunochemical and physiochemical methods, whose major limit is represented by relative low analytical sensitivity. As a consequence, consumers are currently exposed to molecules with potential carcinogenic effects such as 17β-estradiol, the most powerful substance with estrogenic effect. Therefore, high analytical sensitivity screening and conﬁrmatory methods are required, coupling easiness of use and efﬁciency. The aim our activity is developing a high sensitive diagnostic tool intended for monitoring of illegal treatments. We demonstrated the capability of MC biosensors to detect 17β-estradiol in male calves serum: our tool has shown to be able of discriminating treated and untreated animals, showing the ability of detecting traces of 17β-estradiol in serum at concentrations lower than the present accepted physiological serum concentration threshold value (40 ppt) and commercial ELISA tests (25 ppt). The method exhibits a limit of detection of 20 ppt and a limited cross-reactivity with high concentrations (10 ppb) of similar molecules, such as testosterone.
Another important chronic dietary risk factor, more than food additives or pesticide residues, is represented by mycotoxins. Aflatoxins and Ochratoxin A are the most dangerous compounds. The research team showed the feasibility of using microcantilever resonator arrays to effectively identify total aﬂatoxins and ochratoxin A, at low concentrations (3 ng/mL and less than 6 ng/mL, respectively), with relatively low uncertainty (about 10%) and good reproducibility for the same target concentration. Furthermore, the developed immunosensing method shows a limited cross-reactivity to different mycotoxins, paving the way to a highly speciﬁc technique, able to identify different mycotoxins in the sample. To our knowledge, this work represents the ﬁrst example in literature of successfully immunodetection of low concentrations of multiple mycotoxins by microcantilever resonator arrays.
The detection of pathogenic bacteria is the most challenging part of research activity. Two approaches are used to reveal pathogen contamination: in the first one the cantilever are coated with capture antibody able to bind whole cells, the second method considers the detection of pathogen DNA. The detection of Salmonella enterica serotype enteritidis whole cells has been investigated and the experimental limit of detection is lower than 100 cfu/mL.
Gianluca Palmara: PhD student
Alessandro Chiadò, Simone L. Marasso, Stefano Stassi: Post-Doc
Tel. +39 011 090 7398
1. “Development of a microcantilever-based immunosensing method for mycotoxin detection”, C. Ricciardi, R. Castagna, I. Ferrante, F. Frascella, S.L. Marasso, A. Ricci, G. Canavese, A. Lorè, A. Prelle, M.L. Gullino, D. Spadaro. Biosensors and Bioelectronics 40 (2013) 233–239.
2. “Immunodetection of 17β-estradiol in serum at ppt level by microcantilever resonators”,C. Ricciardi, I. Ferrante, R. Castagna, F. Frascella, S.L. Marasso, K. Santoro, M. Gili, D. Pitardi, M. Pezzolato, E. Bozzetta. Biosensors and Bioelectronics 40 (2013) 407–411.
3. “Microcantilver-based DNA hybridization sensors for Salmonella identification”, R. Patti, M.T. Bottero, A. Dalmasso, A. Grassi, I. Ferrante, K. Santoro, N. Ciprianetti, C. Ricciardi. Italian Journal of Food Safety Vol. 1 (2012) 17-19, ISSN: 2239-7132.
4. “Seprion-coated microcantilever sensors for PrPSc detection”, D. Meloni, D. Pitardi, M. Mazza, R. Castagna, I. Ferrante, C. Ricciardi, E. Bozzetta. Prion, 6 (2012) 94-95
5. “Online Portable Microcantilever Biosensors for Salmonella enterica Serotype Enteritidis Detection” C. Ricciardi, G. Canavese, R. Castagna, G. Digregorio, I. Ferrante, S.L. Marasso, A. Ricci, V. Alessandria, K. Rantsiou, L.S. Cocolin. Food Bioprocess Technol 3 (2010) 956–960.