COVALENT BINDING OF PROTEINS TO GRAFTED PLASTIC SURFACES SUITABLE FORIMMUNOASSAYS - II - PICOGRAMS OF IGE DETECTED IN BAL FLUID IN SARCOIDOSIS

An ELISA is described for the quantitation of minute amounts of human IgE using microtiter strips which were surface modified according to a procedure previously described, This ELISA used an IgG fraction of a rabbit polyclonal anti-IgE antibody coupled to the carboxylated surfac e, which prior to coupling had been preactivated with a water soluble carbodiimide. A biotinylated affinity purified rabbit anti-IgE antibod y was uc;ed together with streptavidin conjugated alkaline phosphatase and biotinylated enzyme for the detection of bound analyte. Sarcoidos is is a systemic disorder of as yet unknown cause with epithelioid gra nuloma formation in the lung as a prominent feature. With the bronchoa lveolar lavage technique, epithelial lining fluid was obtained and ana lyzed fur ISE content since differences of immunoglobulin concentratio n in bronchoalveolar lavage fluid (BALF) may reflect disease activity. Our modified ELISA system was used for measuring IgE in BALF from hea lthy controls and sarcoid patients. The assay using modified strips de monstrated a considerably improved sensitivity compared to the use of physical adsorption of first antibody to untreated strips, The detecti on limit of the improved assay was approximately 3 pg IgE/sample (100 mu l). Quantitative recovery of IgE was demonstrated within the measur ing range (51.2-12500 pg IgE/ml) and more than 40% (23/54) of the sarc oid patients showed values above the lowest standard concentration (51 .2 pg/ml). In contrast, none of the healthy controls (0/22) had detect able ISE as defined by the detection limit of the Essay. (C) 1997 Else vier Science B.V.