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  • Title: A nonseparation, time-resolved fluoroimmunoassay to monitor ovarian function and predict potential fertility in women.
    Author: Barnard G, O'Reilly C, Dennis K, Collins W.
    Journal: Fertil Steril; 1989 Jul; 52(1):60-5. PubMed ID: 2744188.
    Abstract:
    The authors describe a nonseparation, time-resolved fluoroimmunoassay involving the use of monoclonal antibodies for the measurement of estrone-3-glucuronide in urine. The method has appropriate sensitivity (12 nmol/l), better specificity than a conventional radioimmunoassay with polyclonal antibodies, and the advantages of speed, simplicity, less imprecision, and improved clinical effectiveness. The labeled antigen is a novel fluorescent europium chelate covalently linked to estrone glucuronide at carbon 6 of the glucuronide moiety. The antibodies were raised against estrone-3-glucuronyl-6-bovine serum albumin. The antibody binding reaction is performed in microtiter wells (or tubes) and involves the addition of labeled antigen in buffer (2 ng/100 microliters), a limited concentration of antibodies in buffer (100 microliters), and standard or urine sample (10 microliters). The mixture is incubated for 15 minutes at room temperature. Time-resolved fluorescence from the unbound label is proportional to the concentration of estrone glucuronide. The method may be used to monitor ovarian function and potential fertility in women. In the study and measurement of analytes which indicate the status of reproduction, immunoassays are still being updated and utilized. Nonseparation immunoassays have been specifically developed in the study of drugs in serum or urine and most likely can be developed for the measure of urinary estrone glucuronide. The advantages of this process include accuracy, simplicity, speed and the possibility of full automation. However, nonseparation immunoassays are affected by nonspecific interference from the attributes within biologic materials, assay buffers, reagents and plastics. The use of time-resolved fluorescence has been implemented and has been proven to be successful. The variety of assays used in the measurement of haptens and large molecular weight analytes has been greatly improved. The technique of using time-resolved fluorescence gives us the greatest potential in the production of the most sensitive separation and nonseparation immunoassays. This method has more appropriate sensitivity and a better specificity than typical radioimmunoassays with polyclonal antibodies. This process may be used in the study of ovarian functions and future fertility in women.
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