FOOD SAFETY SUPPLEMENT: RAPID METHODS 05


interactions of interest in food chemistry are matrix and processing effects and cross- reactivity to similar analytes. Processing effects and food matrixes can


have a significant impact on the ability of an ELISA to detect the analyte of interest. In a recent study it was shown that roasting peanuts had a significant decrease in the detection of allergenic proteins and the ability to determine peanut content15


. This could be due to changes in the


solubility, protein structure or changes in the epitopes during processing. In fact, any change in the analyte, such as the masking or conjugation of mycotoxins16


, will compromise


the detectably in immunological assays. Components in the matrix can also have a


» The occurrence of allergens in food is another important topic in the area of food safety «


significant impact on the performance of the assay. For example it is well known that chocolate contains components that interfere with the detection of allergens and require special extraction cocktails17


, while components in


coffee can produce overestimates in Ochratoxin A assays and require additional steps to remove these matrix components from solution13


. Finally, the specificity of the immunological


assay is an important aspect that needs to be considered. Generally speaking, antibodies are very specific to the target analyte, but well validated methods will determine the cross- reactivity of the assay to potential components in the food or other chemicals with similar structures to the target analyte. Although cross- reactivity is typically an unwanted characteristic in an ELISA, it can have an advantage of determining total contamination to a class of compounds18


. Well validated immunological methods are


very powerful and have a great potential as rapid screening tools for food safety, but there are caveats to using these methods and they must be kept in mind when analysing samples or deviating from well validated protocols.


PCR methods In the past three decades, novel culture- independent approaches have undergone considerable development for studying microbial populations in food. Among these, the PCR based techniques are considered as the


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method of choice for the detection and quantification of microorganisms19


. The flexible


PCR system allows either the specific detection of one particular strain in the background of other organisms or of a whole group of strains or even genera sharing a common feature can be detected. Besides the high specificity of PCR tests, the sophisticated sensitivity of these assays – even a single DNA molecule can be detected – helped to get this technique widely accepted for food analysis. The PCR reaction is used to amplify even minor amounts of DNA to a detectable level. While in the early years of the PCR visual systems like dye stained agarose gels were used for qualitative product detection, nowadays modern fluorescence detection systems are applied. These novel technologies allow the real-time monitoring of the amplification of target DNA and thus enable quantification. Furthermore, up-to-date PCR machines can be used to measure up to six analytes in a single reaction and depending on the machine, 96 or 384 reactions can be run in approximately 45 minutes. These benefits allow the integration of the PCR in high throughput analysis cascades in which the reaction set-up is performed by automated liquid handling stations. PCR and quantitative PCR (qPCR) are now well established for the analysis of food samples, especially for the detection of foodborne pathogens or genetically modified organisms19,20


. Apart from these important fields of PCR


application, this technique can also be used to screen samples for potential mycotoxin producing fungi. Numerous quantitative PCR tests have been developed to measure the amount of DNA of mycotoxigenic strains belonging to the genera Fusarium, Aspergillus, and Penicillium21


. However, targeting single


strains is too laborious to be applied for practical studies as all samples have to be checked for each potentially occurring strain. To over - come this drawback, modern qPCR tests focus on the key genes of biosythetic pathways. Usually, the gene encoding for an enzyme that catalyses the formation of the starting metabolite of a mycotoxin synthesis pathway gives a perfect target. All organisms sharing this gene have the potential to produce a certain class of mycotoxins, independent of their phylogenetic origin. The first PCR assays for mycotoxin


producing fungi were published in the 1990s with the aim to detect aflatoxin producers22,23 Since then numerous variations of the


. New Food Volume 14, Issue 5, 2011


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