Identification of peanut and hazelnut allergens by native two-dimensional gel electrophoresis

Hird, Heather; Pumphrey, R. S. H.; Wilson, Philip B.; Sunderland, Joanne; Reece, Paul

doi:10.1002/1522-2683(20000701)21:13<2678::aid-elps2678>3.0.co;2-v

Cited by 28 publications

(11 citation statements)

References 12 publications

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“…As in any other immunoanalytical application, cross reactivity has to be critically investigated. Precipitation of the proteins with organic solvents, washing, lyophilisation, and extraction with aqueous buffer are the protein-purification techniques most commonly used in analytical laboratories [12,13,14,15]. Such additional purification steps depend largely on the aim of the analysis -if any of the soluble proteins present in the extract of one commodity are to be detected ("protein-assay"), no further isolation steps are usually necessary during a immunoassay development.…”

Section: Immunoanalytical Techniquesmentioning

confidence: 99%

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Immunoanalytical detection of allergenic proteins in food

Krska¹,

Welzig²,

Baumgartner³

2004

Analytical and Bioanalytical Chemistry

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Section: Immunoanalytical Techniquesmentioning

confidence: 99%

“…1 Developing "allergen" and "protein" immunoassays for determination of allergenic proteins in food (example for peanuts is given). Pabpolyclonal antibodies, mab -monoclonal antibodies and RIE have all been found suitable for separating allergenic proteins [14,16,17,18]. Among these, SDS-PAGE is most frequently used.…”

Section: Immunoanalytical Techniquesmentioning

confidence: 99%

Immunoanalytical detection of allergenic proteins in food

Krska¹,

Welzig²,

Baumgartner³

2004

Analytical and Bioanalytical Chemistry

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“…At present, different techniques, such as immunoassays, immunoblotting, immuno-electrophoresis, real-time polymerase chain reaction (PCR), PCR-enzyme-linked immunosorbent assay (ELISA) and mass-spectrometry are available to detect and/or quantify (trace amounts of) hazelnut proteins (Kiening et al 2005, Ben Rejeb et al 2003, Arlorio et al 2007, Hird et al 2000, van Hengel 2007, Blais and Phillippe 2001, Yman et al 2006, Scheibe et al 2001, Holzhauser et al 2002. However, the value of the results obtained with these techniques will strongly depend on the quality of the extract used.…”

Section: Introductionmentioning

confidence: 99%

Comparison and Functional Evaluation of the Allergenicity of Different Hazelnut (Corylus avellana) Protein Extracts

et al. 2010

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This paper investigates two methodological issues of hazelnut protein extraction: the use of protease inhibitors during protein extraction and the effect of defatting hazelnuts before protein extraction. Different protein extracts from hazelnuts were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and IgE-immunoblotting to evaluate the presence of allergens in the extract. The allergy-provoking potential of these extracts was assessed by the Basophil Activation Test. This functional test relies on in vitro stimulation of human basophils from hazelnut-allergic patients with the different extracts. SDS-PAGE and IgE-immunoblot analysis of hazelnut extracts prepared without protease inhibitors revealed partial degradation of the proteins in the extracts. This observation was confirmed by the Basophil Activation Test, as the percentage of activated basophils was lower as compared to extracts prepared with protease inhibitors. Biochemical and functional evaluation of a defatted hazelnut extract did not reveal differences in comparison with non-defatted extracts. This study is the first to report on the use of the Basophil Activation Test applying human basophils for functional evaluation of protein extracts from hazelnuts. The results demonstrate that the use of protease inhibitors during extraction is essential for the stability and functionality of a protein extract, whereas defatting does not affect the quality of the protein extract.

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“…Figure 7 shows the production of TNF¬ as a result of sensitization with two types of peanut preparations, one from a skin prick formulation and one an FPLC puri®ed peanut fraction. The skin prick formulation shows very little activity above control 1104 S. Oehlschlager et al Hird et al 2000 values, in contrast to the puri®ed fraction. Secondly there is considerable variation in the response of the lung tissues, and the system cannot be considered robust.…”

Section: Approachesmentioning

confidence: 87%

“…Cross-reactivit y can be examined experimentally using native 2-D gel electrophoresis (Hird et al 2000). In this approach proteins retain epitopic structure following separation and can be probed with IgE-bearing sera.…”

Section: Approachesmentioning

confidence: 99%

Food allergy—towards predictive testing for novel foods

Oehlschlager¹,

Reece²,

Brown³

et al. 2001

Food Additives and Contaminants

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The risks associated with IgE-mediated food allergy highlight the need for methods to screen for potential food allergens. Clinical and immunological tests are available for the diagnosis of food allergy to known food allergens, but this does not extend to the evaluation, or prediction of allergenicity in novel foods. This category, includes foods produced using novel processes genetically modified (GM) foods, and foods that might be used as alternatives to traditional foods. Through the collation and analysis of the protein sequences of known allergens and their epitopes, it is possible to identify related groups which correlate with observed clinical cross-reactivities. 3-D modelling extends the use of sequence data and can be used to display eptiopes on the surface of a molecule. Experimental models support sequence analysis and 3-D modelling. Observed cross-reactivities can be examined by Western blots prepared from native 2-D gels of a whole food preparation (e.g. hazelnut, peanut), and common proteins identified. IgEs to novel proteins can be raised in Brown Norway rat (a high IgE responder strain) and the proteins tested in simulated digest to determine epitope stability. Using the CSL serum bank, epitope binding can be examined through the ability of an allergen to cross-link the high affinity IgE receptor and thereby release mediators using in vitro cell-based models. This range of methods, in combination with data mining, provides a variety of screening options for testing the potential of a novel food to be allergenic, which does not involve prior exposure to the consumer.

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Identification of peanut and hazelnut allergens by native two-dimensional gel electrophoresis

Cited by 28 publications

References 12 publications

Immunoanalytical detection of allergenic proteins in food

Immunoanalytical detection of allergenic proteins in food

Comparison and Functional Evaluation of the Allergenicity of Different Hazelnut (Corylus avellana) Protein Extracts

Food allergy—towards predictive testing for novel foods

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