As part of our Integrated and Standardised Global Quality Program, Mérieux NutriSciences Corporate Chemistry and Quality teams have decided that all Mérieux NutriSciences network of laboratories use only the MORINAGA Egg (Ovalbumin) ELISA Kit II for egg analysis.
The new Limitation of Quantification (LOQ) MORINAGA (Ovalbumin ) ELISA is 1 ppm.
This change will apply on Monday 24 December.
What are the main changes for Egg analysis?
The MORINAGA (Ovalbumin) ELISA kit will be used for Egg analysis (Silliker method M 133).
There is evidence that shows that several egg ELISA kits are not able to detect some egg allergenic proteins in processed food products. Several scientific papers explain (see the references below) explain that an improved procedure that includes reducing agents needs to be used to efficiently extract the egg allergenic proteins in order to detect them.
The ELISA Egg kit from the Morinaga company is based on this principle, and it was demonstrated to be the most efficient kit for the detection of egg proteins in many processed food (bakery, pasta, cooked food). The accuracy of this kit has been further confirmed by internal validation work conducted by Mérieux NutriSciences laboratories.
Consequently, Mérieux NutriSciences Corporate Chemistry and Quality team have decided that all Mérieux NutriSciences network of laboratories use only the MORINAGA Egg (Ovalbumin) ELISA Kit II for food analysis.
Qualitative and quantitative differences can be expected when comparing Morinaga Egg kit results with kits from other brands, including Proficiency Tests and Interlaboratory trials. This could sometimes puzzle customers about result interpretation, and consequent actions to be taken.
But, as an important carrier for the safety and protection of human health, Mérieux NutriSciences can only decide on the application of the best available Science, in this particular case represented by Morinaga kit.
- Extractability, stability, and allergenicity of egg white proteins in differently heat processed foods, Faeste CK, et al, J AOAC Int. 2007;90:427–36
- Multi-allergen Quantitation and the Impact of Thermal Treatment in Industry-Processed Baked Goods by ELISA and Liquid Chromatography-Tandem Mass Spectrometry, Parker et al, J Agric Food Chem. 2015 Dec 16;63
- Food allergen analysis for processed food using a novel extraction method to eliminate harmful reagents for both ELISA and lateral-flow tests, Kaori et al, Analytical and Bioanalytical Chemistry, 408, 5973-5984 (2016)
- Evaluation and Validation of a Commercial ELISA-Based Method for the Determination of Egg Protein in Foods, Eric Marceau et al, Canadian Food Inspection Agency study (2013 poster)
- Evaluation of Commercial ELISA Assays for the Detection of Egg in Food, Garber E. et al, Division of Natural Products, OPDF, CFSAN, U.S. Food and Drug Administration (poster)
Multi-allergen Quantitation and the Impact of Thermal Treatment in Industry-Processed Baked Goods by ELISA and Liquid Chromatography-Tandem Mass Spectrometry. Parker et al, J Agric Food Chem. 2015 Dec 16;63
Undeclared food allergens account for 30−40% of food recalls in the United States. Compliance with ingredient labeling regulations and the implementation of effective manufacturing allergen control plans require the use of reliable methods for allergen detection and quantitation in complex food products. The objectives of this work were to (1) produce industry processed model foods incurred with egg, milk, and peanut allergens, (2) compare analytical method performance for allergen quantitation in thermally processed bakery products, and (3) determine the effects of thermal treatment on allergen detection. Control and allergen-incurred cereal bars and muffins were formulated in a pilot-scale industry processing facility. Quantitation of egg, milk, and peanut in incurred baked goods was compared at various processing stages using commercial enzyme-linked immunosorbent assay (ELISA) kits and a novel multi-allergen liquid chromatography (LC)-tandem mass spectrometry (MS/MS) multiple-reaction monitoring (MRM) method. Thermal processing was determined to negatively affect the recovery and quantitation of egg, milk, and peanut to different extents depending on the allergen, matrix, and analytical test method. The Morinaga ELISA and LC-MS/MS quantitative methods reported the highest recovery across all monitored allergens, whereas the ELISA Systems, Neogen BioKits, Neogen Veratox, and R-Biopharm ELISA Kits underperformed in the determination of allergen content of industry-processed bakery products.
Comparison of commercial ELISA kit performance for whole egg powder in 5000 ppm incurred (A) cereal bar and (B) muffin samples collected at the mixer (blue) and final product (red).
Measured concentrations are normalized to ppm (μg/g) allergen protein and corrected for moisture content.
The dotted line in each graph represents allergen recovery at 100%.
Method performance as influenced by heat processing is reported above each ELISA kit as a ratio of measured allergen protein concentration in the final product and mixer samples.
Extractability, stability, and allergenicity of egg white proteins in differently heat processed foods. Faeste CK et al, J AOAC Int. 2007;90:427–36
Hen's egg white protein is a major cause of food allergy, and a considerable number of countries have introduced labeling directions for processed food products. To control compliance with these regulations, analytical assays for the detection of egg in manufactured foods have been developed. In this study, we have tested the performance of 3 commercially available kits for quantitative egg analysis using 6 model heat-processed foods. The 3 assays worked well under standard conditions with soluble egg white proteins, but only the kit using a denaturing-reducing extraction buffer detected egg in complex heat-treated food matrixes. The differently extracted food samples were further used to evaluate the stability and allergenicity of the egg white allergens ovalbumin, ovomucoid, ovotransferrin, and lysozyme with polyclonal anti-egg antibodies and sera of 6 patients with egg allergy. It could be shown that differences in egg protein extractability have a significant impact on the interpretation of study results.
Evaluation and Validation of a Commercial ELISA-Based Method for the Determination of Egg Protein in Foods, Eric Marceau et al, CFIA study 2013
The detection of food allergens like egg protein in heat processed foods using ELISA-based methods has always been challenging. The Canadian Food Inspection Agency has evaluated the performance of the Morinaga Institute of Biological Science ELISA Egg Kit through an inter-laboratory study. Use of the method improves the probability to detect egg allergen in processed products, which will provide further protection to egg-allergic consumers. After a successful single laboratory validation where a conversion factor was established for the NIST 8445 material, a total of 124 samples were distributed to four laboratories. Each participant received 10 ice cream, 10 bread and 11 pasta samples. Each food commodity was either egg free, already contained egg at low (about 4 ppm), or high (about 1800 ppm) level or was spiked at a level of 1.7, 6.7 or 16.8 ppm using the carboxymethylcellulose suspension technique. Z-Scores were calculated for the incurred positive samples and were all well within the acceptable range. The precision (RSDR) ranged from 5.1 to 12.4% for these materials. The results for spiked samples were close to the expected values and the recoveries varied by commodity, with the ice cream averaging 99%, the bread 101% and the pasta 107% for an overall recovery of 103%. No false negatives were found while a single false positive result was reported. Overall the validation data were deemed satisfactory and confirmed the fitness-for-purpose of the method.
Food allergen analysis for processed food using a novel extraction method to eliminate harmful reagents for both ELISA and lateral-flow tests. Kaori et al, Analytical and Bioanalytical Chemistry, 408, 5973-5984 (2016)
Enzyme-linked immunosorbent assay (ELISA) is commonly used to determine food allergens in food products. However, a significant number of ELISAs give an erroneous result, especially when applied to highly processed food. Accordingly, an improved ELISA, which utilizes an extraction solution comprising the surfactant sodium lauryl sulfate (SDS) and reductant 2-mercaptoethanol (2-ME), has been specially developed to analyze food allergens in highly processed food by enhancing analyte protein extraction. Recently, however, the use of 2-ME has become undesirable. In the present study, a new extraction solution containing a human- and eco-friendly reductant, which is convenient to use at the food manufacturing site, has been established. Among three chemicals with different reducing properties, sodium sulfite, tris(3-hydroxypropyl)phosphine, and mercaptoethylamine sodium sulfite was selected as a 2-ME substitute. […] The ELISA based on the SDS/0.1 M sulfite extraction solution has now been authorized as the revised official method for food allergen analysis in Japan.
Edit note: Morinaga uses SDS/2-ME (now dismissing) and SDS/Sodium Sulfite, while most diffused competitors use SDS.
Evaluation of Commercial ELISA Assays for the Detection of Egg in Food, Garber E. et al, Division of Natural Products, OPDF, CFSAN, U.S. Food and Drug Administration (poster)
It is estimated that 6% of children under the age of 3 are allergic to some form of food with 1.3% allergic to eggs. Currently, the only method available for individuals allergic to a specific food to avoid an allergic reaction is avoidance of the allergenic food. To address the need for validated methods to test food products for the presence of allergens, the FDA has initiated studies to evaluate commercial immunology based assays for the detection of allergenic foods. Seven commercial ELISA-based assays for the detection of egg proteins were evaluated with six different matrices representative of various forms of processing. The matrices examined included baked goods, pasta - analyzed before and after boiling, vanilla ice cream, salad dressing (no processing), and phosphate buffered saline. Each of the matrices were spiked with either 0, 2, 5, 10, 25, or 100 µg/g of the NIST whole egg powder standard reference material (SRM) # 8415. Six of the ELISA kits relied on washing/partitioning to extract the antigenic biomarker into an aqueous buffer. The seventh kit was unique in that it employed reducing–denaturing conditions to extract the antigenic proteins. All seven kits readily detected egg spiked into foods which were subjected to none or minimal heating. However, the assays displayed significant differences in ability to detect egg in foods that were exposed to heat. Only the kit that employed reducing-denaturing conditions to extract the antigenic biomarkers detected egg in all matrices spiked with > 2 µg/g of the NIST SRM.
Taylor SL, Nordlee JA, Niemann LM, Lambrecht DM. Allergen immunoassays – considerations for use of naturally incurred standards. Anal Bioanal Chem. 2009;395:83–92.
The enzyme-linked immunosorbent assay (ELISA) offers many advantages for the detection of potentially hazardous allergenic food residues that might become adventitious components of other foods during the course of food production and processing. ELISAs detect proteins, and food allergens are proteins. ELISAs are sufficiently sensitive and specific for detection of food allergen residues. ELISAs can also be produced in formats that are compatible with the industrial food processing environment. However, ELISAs also have disadvantages that should be carefully evaluated and widely recognized. Various food-processing operations can have profound effects on the detectability of allergenic food residues. ELISAs detect intact proteins but protein hydrolysates evade detection in some ELISA formats. The residual proteins present in some ingredients derived from commonly allergenic sources may also not be easily detected with ELISAs because of the nature of the protein residues remaining, e.g. lipophilic. Processing operations can dramatically lower the solubility of proteins. In some food formulations, heat processing, in particular, induces chemical modifications that can affect antibody binding to epitopes in the ELISA. The use of naturally incurred standards where allergenic food residues are incorporated into various representative food matrices and then processed in a manner similar to “real-world” food processing can reveal some of the limitations of allergen ELISAs. Methods for the preparation of naturally incurred standards in chocolate, cookie, muffin, ice cream, pasta, frankfurter, and cream of potato soup are provided as examples.
Effect of Heat Treatment on the Quantitative Detection of Egg Protein Residues by Commercial Enzyme-Linked Immunosorbent Assay Test Kits, Tong-Jen Fu et al, J. Agric. Food Chem., 2010, 58 (8), pp 4831–4838
This study examined the changes in the solubility of egg proteins as affected by different heat treatments and compared the performances of three commercial test kits for the quantitation of protein residues in heat-treated samples. National Institute of Standards and Technology (NIST) whole egg standard reference material #8415 and Henningsen spray-dried whole egg powder were subjected to heating in the presence of water at 60 and 100°C, autoclaving for 5 or 10 min, or dry heating at 60−400°C for 10 min. The amount of protein in the heated samples was assayed using the bicinchoninic acid total protein assay as well as egg-specific commercial enzyme-linked immunosorbent assay (ELISA) kits. Elevated heat resulted in a lower level of proteins extracted. Neogen's Veratox kit, which is reactive to multiple proteins in egg, greatly underestimated the amount of residual proteins in the boiled or autoclaved samples. Tepnel BioSystems' Biokits assay, which employs antibodies specific to a heat-stable marker protein (ovomucoid), registered a higher level of protein in these samples. Both test kits substantially underestimated the amount of residual proteins in samples dry-heated at temperatures >176 °C. The Morinaga test, using an improved extraction buffer, registered the highest level of protein in the heat-treated NIST samples but not the Henningsen samples. The underestimation by the commercial test kits was attributed to changes in the immunoreactivity of residual proteins after heat treatments and not the differences in the amount of protein extracted. These results suggest that thermal processing may affect the quantitative analysis of allergens and needs to be taken into account in the validation of commercial ELISA test kits.