Development of a functional in vitro assay as a novel tool for the standardization of allergen extracts in the human system.
BACKGROUND
Biochemical and immunochemical methods used for batch control of allergen extracts rely on the binding of IgE molecules to allergens. They do not measure the ability of a protein to induce type I allergic reactions. Therefore, a biological assay was established that is based on the cellular mechanisms of allergies in order to assess the cross-linking capacity of allergens.


METHODS
Rat basophilic leukaemia cells were transfected with cDNA coding for the human high affinity IgE receptor chains. The surface expression of the IgE-binding alpha-chain was detected by FACS analysis and the functional integration of the 'humanized' receptors into the signal transduction cascade was addressed by intracellular calcium mobilization. Mediator release was measured in response to human IgE and a variety of cross-linking allergen preparations.


RESULTS
Several clones were obtained that were able to bind allergen-specific human IgE. The results of the biological assay were compared with those obtained by immunochemical methods. The biological assay was used to determine the potency of allergen extracts, including highly diluted products that cannot be analysed by conventional methods.


CONCLUSIONS
A stable 'humanized' basophil cell line was established that will be a useful tool for the standardization and batch control of allergen extracts. Because of its high sensitivity, it can also be used to detect minute quantities of potentially allergenic proteins, e.g. in processed foods. In addition, the test may support the development of novel allergy vaccines, such as recombinant hypoallergenic molecules.
Vogel
L
L
Department of Allergology, Paul-Ehrlich-Institut, Langen, Germany.
Lüttkopf
D
D
Hatahet
L
L
Haustein
D
D
Vieths
S
S
D000485:Allergens / Q000032:analysis* / Q000276:immunology; D000818:Animals; D000918:Antibody Specificity; D001491:Basophils / Q000276:immunology; D001681:Biological Assay; D002460:Cell Line; D018076:DNA, Complementary; D006801:Humans; D007150:Immunohistochemistry; D007158:Immunologic Techniques; D020033:Protein Isoforms / Q000276:immunology; D051381:Rats; D017455:Receptors, IgE / Q000235:genetics / Q000276:immunology; D012015:Reference Standards; D014162:Transfection
D000485:Allergens; D018076:DNA, Complementary; D020033:Protein Isoforms; D017455:Receptors, IgE