Characterization of IgG-subclass antibodies to individual allergens by crossed radioimmunoelectrophoresis.

A crossed radioimmunoelectrophoretic (CRIE) method for detection of human IgG subclass specificities against individual antigens is described. After crossed immunoelectrophoresis (CIE) of the antigens the immunoplates are incubated with serum, washed and incubated with mouse monoclonal anti-human IgG-subclass antibodies. After washing, the plates are finally incubated with the 125I-labelled detector protein, rabbit anti-mouse IgG, and washed. The plates are then placed on an X-ray film for autoradiography. The specificity of the method was tested by inhibition with antigens in the first layer and by inhibition with myeloma sera containing only one IgG subclass protein in the second layer. The specificity of the third layer was assured by affinity purification of the rabbit anti-mouse IgG antibodies. The method was shown to be sensitive and specific. The test systems were timothy (Phleum pratense) pollen antigens and house dust mite (Dermatophagoides pteronyssinus) antigens.