Detection of harmful epitopes in foods - Extraction: the Achilles heel of allergen testing
9 important questions on Detection of harmful epitopes in foods - Extraction: the Achilles heel of allergen testing
Why is a correct extraction procedure of allergens important?
Which factors control the successful extraction of the allergen?
- Solubility of the allergen in the extraction solvent
- Pre-treatment of the matrix
- Competing solvents/binding to the matrix
- Effect of the extraction on the conformational epitopes
Mention 3 criteria for extraction method
- The method should allow complete extraction of the allergen
- The extraction protocol should work independent of the food matrix analysed
- The extraction method should be compatible with the detection method used
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The allergen needs to be soluble in the extracting. This can cause some problems:
Why are gluten proteins difficult to extract? How can this be solved?
- Other extraction temperatures can be used.
- Another buffer system
- Higher salt concentrations, if the test is sufficiently sensitive and they can be 'diluted out' (dilutions > 100x)
What can be done to increase the amount of allergen extracted from the sample?
- Different type of buffer
- Presence of salts
What can be done to make sure extraction procedures extract all proteins (also the denatured ones)?
Why is the situation more complex, if you want to test for allergens in a food matrix?
- The antibodies used must be able to react with the allergen. An antibody against a conformational epitope is unwanted in this type of assays.
- The allergen must be soluble in the extraction solvent. For some proteins to become soluble, the disulphide bonds have to be reduced. Under these conditions, antibodies themselves are reduced and lose their ability to interact.
- Even if the antigen is soluble, the matrix may not. The antigen is entrapped in the matrix and can only be extracted after disintegration of the matrix. Pre-processing of the matrix by grinding or milling is always a requirement.
Why is extraction of peanut allergen in dark chocolate difficult?
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