| abstract |
The invention pertains to a method for the determination of basophil activation induced by a test substance by flow cytometric measurement of the changes of the mean or median fluorescence intensities (MFI) of the basophilic F c εRI receptor present on the cell surface of basophils (MFI-F c εRI) and/or the IgE antibodies bound to the F c εRI receptor (MFI-IgE), and the CD63 antigen exposed on the cell surface of basophils after their activation (MFI-CD63), by means of a mixture of anti-CD63, anti-F c εRI or anti-IgE, and anti-CCR3 antibodies each labelled with a distinct fluorophore, of which at least one antibody acts as a basophil selection marker and at least two antibodies act as basophil activation markers, and bringing the mean fluorescence intensities of the activation markers in correlation to obtain an Activation Index. These methods combining the measurement of an early (such as IgE, F c εRI or CD203c) and a late basophil activation marker (such as CD63), respectively, provide a markedly improved clinical sensitivity in allergy diagnosis over existing methods which consider only one activation marker, such as CD203c or CD63, expressed in percentage of basophil activation. It is also an aspect of the present invention to provide an ex-vivo allergy provocation test comprising the above-mentioned flow cytometric measurement and analysis of the results as well as a test kit for carrying out the test in-vitro. |