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This document provides supplemental guidance on specifications for the development and implementation of studies to validate the performance characteristics of quantitative ELISA methods for the determination of food allergens. It is intended as a companion document to other existing publications on method validation. The guidance is divided into two sections: information to be provided by the method developer on various characteristics of the method, and implementation of a multilaboratory validation study. Certain criteria included in the guidance are allergen-specific. Two food allergens, egg and milk, are used to demonstrate the criteria guidance. These recommendations will be the basis of the harmonized validation protocol for any food allergen ELISA method, whether proprietary or nonproprietary, that will be submitted to AOAC and/or regulatory authorities or other bodies for status recognition. Regulatory authorities may have their own particular requirements for data packages in addition to the guidance in this document. Future work planned for the implementation and validation of this guidance will include guidance specific to other priority allergens.

The objective of this study was to evaluate the hypothesis that the dose-response relationship for Listeria monocytogenes in humans varies with genotypic lineage or subtype. The linkages between molecular subtyping data and enumeration data for L. monocytogenes subtypes in foods consumed by the at-risk population were examined to test this hypothesis. We applied a conditional probability model to conduct a subtype-specific dose-response analysis, with the focus on invasive listeriosis. L. monocytogenes differed not only in the molecular subtype and lineage but also in the contamination level when isolates of the pathogen occurred in retail samples of ready-to-eat foods. Using the exponential model parameter r-value as a measure (essentially the probability of a single cell causing illness), we found that the virulence varied among L. monocytogenes lineages by several orders of magnitude. Under the assumptions made, for L. monocytogenes lineages I and II the consumption of a single cell would result in listeriosis with log average probabilities of -7.88 (equivalent to once in 10(7.78) times) and -10.3, respectively, as compared with -9.72 for L. monocytogenes independent of subtype. A greater difference in r-values was found for selected ribotypes. The uncertainty about the r-value estimates was small compared with the large differences in the virulence parameters themselves. Thus, for L. monocytogenes both subtype and the number of cells consumed matter, highlighting the usefulness of considering both exposure concentration and subtype prevalence in dose-response analysis. As advances are made in molecular subtyping and quantitative tools for dose-response analysis, further studies integrating genomic data into quantitative risk assessments will enable better attribution of disease risk to L. monocytogenes subtypes.

Because of the public health significance of L. monocytogenes, U.S. regulatory agencies established a policy whereby ready-to-eat foods contaminated with the organism at a detectable level are deemed adulterated. This "zero tolerance" policy, however, makes no distinction between foods contaminated at high and low levels. We have reported elsewhere that a survey of over 31,000 ready-to-eat retail food samples, representing eight product categories, showed an overall prevalence rate of 1.82% for these foods. In this study, we used the food survey data in combination with concurrent data regarding illness in the population consuming the foods, together with other variable factors, to derive a dose-response model. The confidence interval for prevalence was 1.68 to 1.97%. L. monocytogenes levels, which ranged from -2 to 6 log CFU/g, were adequately described by the distribution beta (0.29, 2.68, -1.69, 6.1). An exponential dose-response model was obtained, with an R value (essentially the probability of a single cell causing illness) of 1.76 x 10(-10) for the population at the highest risk. A microbial risk assessment based on the model shows that an alternative to the zero tolerance strategy has a greater risk reduction potential and suggests that a management strategy focusing on the concentration of L. monocytogenes rather than its presence alone may have a greater impact on the improvement of public health by facilitating the development of control measures to limit the maximum levels of L. monocytogenes in foods.