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Iodinated versus fluorescent labelling in the radioallergosorbent test (RAST) for the determination of serum IgE levels

Journal of Immunological Methods
Publication Date
DOI: 10.1016/0022-1759(76)90108-3
  • Biology


Abstract Experiments were performed to determine the feasibility of replacing the radioactive 125I tag in the radioallergosorbent test (RAST) with a new fluorescent compound, 2-methoxy-2, 4-diphenyl-3 (2H)-furanone (MDPF). It was possible to discriminate a grassallergic from a grass-non-allergic patient using the traditional RAST procedure with 125I-anti-IgE. However, when a fluorescent label, MDPF was attached to IgE using the same RAST procedure, it was not possible to discriminate between allergic and non-allergic responses. Additional experiments, using BSA and anti-BSA as a model, showed that it was possible to detect 0.025 ppm antigen using iodinated antiserum while the fluorescent tag had a lower limit of antigen detection of 0.20 ppm. It was possible to distinguish between allergic and non-allergic responses using 125I-N-succinimidyl-3(4-hydroxyphenyl) propionate (SHPP) which binds to protein in a manner similar to MDPF. These results suggested that the unsatisfactory fluorescence results were not due to an alteration of protein structure as a result of MDPF labelling. Although the fluorescence work was performed using a fluorimeter with optics and electronics optimized for detection of MDPF, sensitivity of the method was limited by high fluorescent background.

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