Quantitation of heat-treated egg with commercial ELISA kits
A study conducted by the US FDA has examined the impact of different heat treatments on egg proteins, and compared the performances of three commercial test kits for the quantitation of egg protein residues in the heat-treated samples.
The samples included in the study were: National Institute of Standards and Technology (NIST) whole egg standard reference material #8415 and Henningsen spray-dried whole egg powder. The samples were heated in the presence of water at 60 and 100°C, autoclaved for 5 or 10 min, or subjected to dry heating at 60-400°C for 10 minutes.
The amount of protein in the heated samples was assayed using the bicinchoninic acid total protein assay, and with the egg-specific ELISA kits commercially available from Neogen, Tepnel and Morinaga.
The results reported in the study are as follows: Elevated heat resulted in a lower level of proteins being extracted. The Neogen Veratox kit, which is reactive to multiple proteins in egg, greatly underestimated the amount of residual proteins in the boiled or autoclaved samples. The 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 the Neogen and Tepnel 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 each of the commercial test kits used in this study was attributed to changes in the immunoreactivity of residual proteins after heat treatments and not the differences in the amount of protein extracted.
The authors conclude that thermal processing impacts on the quantitative analysis of allergens and suggest this factor needs to be taken into account in the validation of commercial ELISA test kits.
Reference: Fu et al. 2010 Journal of Agricultural and Food Chemistry. Vol 58(8) pp. 4831-8. DOI: 10.1021/jf903536z.
