Application of microtitre plates and fluorescence reading to shorten handling of Phadezym RAST and Phadezym IgE PRIST.
To enable shorter and more convenient testing, the Phadezym RAST and Phadezym IgE PRIST procedures for the determination of specific and total IgE were modified in three ways: (i) allergen-coupled paper discs were tested in microtitre wells; (ii) the incubation times were reduced to 1 hr with serum and 2 hr with the anti-IgE by shaking the plates at room temperature; and (iii) the fluorogenic substrate used reduced the development time to 15 min. Determination of IgE antibody specific for fifteen inhalation allergens by the modified fluorescence test (FEIA) and by the conventional Phadezym RAST (EIA) was performed on the serum of thirty-two patients suffering from asthma/rhinitis: correlation studies for these sera showed that 96.1% of the results fell in the same class. In these patients, both FEIA and EIA detected the same proportion of skin-prick tests (SPT) positive results (67%). With the FEIA, 4/165 (2.4%) class 1 results were found in eleven non-atopic subjects (symptom free, fifteen negative SPT, total IgE lower than 80 kU/l), compared to 1/165 (0.6%) with the EIA. In twenty cord sera, both FEIA and EIA found 4/300 (1.3%) class 1 results. For the determination of total serum IgE, the microtitre FEIA showed a detection limit of 0.5 kU/l and an excellent correlation with Phadezym IgE PRIST (n = 66 serum, r = 0.99). These data indicate that the adaptation of Phadezym RAST and Phadezym IgE PRIST to microtitre plates and fluorescence technology has resulted in a time-saving and easy to perform within-day assay which provided results as reproducible, sensitive and specific as those of the conventional procedure.
D000293:Adolescent; D000328:Adult; D001249:Asthma / Q000175:diagnosis; D002648:Child; D005069:Evaluation Studies as Topic; D005260:Female; D005456:Fluorescent Dyes; D005697:Galactosides; D006801:Humans; D006923:Hymecromone / Q000031:analogs & derivatives; D007073:Immunoglobulin E / Q000032:analysis; D008297:Male; D008875:Middle Aged; D011852:Radioallergosorbent Test / Q000295:instrumentation / Q000379:methods*; D011863:Radioimmunoassay / Q000379:methods*; D011864:Radioimmunosorbent Test / Q000295:instrumentation / Q000379:methods*; D012220:Rhinitis / Q000175:diagnosis; D012882:Skin Tests
D005456:Fluorescent Dyes; D005697:Galactosides; D007073:Immunoglobulin E; D006923:Hymecromone; C026845:4-methylumbelliferyl-galactopyranoside