Using evanescent wave immunosensors, a fluorescent labeled analyte may be concentrated within the active surface region by a combination of diffusion to the fiber surface and trapping of the molecule or complex by an antigen-antibody affinity reaction. With the dye B- phycoerythrin, approximately 1.5 X 10-22 moles may be sensed over a 1 mm2 surface. From this number (as determined for a particular dye and instrumentation system) and antibody affinity constants, limits to assay sensitivity can be calculated and the kinetics modeled.
The principles of evanescent wave theory were applied to an immunological sensor for detecting the cardiac-specific isoenzyme creatine kinase-MB (CK-MB). The detection of the CK-MB isoenzyme is used in conjunction with the total CK measurement in the diagnosis of acute myocardial infarction. The clinical range for CK-MB is from 2-100 ng/ml. Previous work which utilized the fluorophor, Fluorescein isothiocyanate (FITC), was able to discriminate between 0 and 3 ng/ml CK-MB. Use of the fluorophor B-phycoerythrin (BPE) increased the assay sensitivity to 0.1 ng/ml CK-MB. The data was collected for 15 minutes using an optical launch and collection angle of 25 degree(s). This fiber optic based system is homogeneous and requires no subsequent washing, handling, or processing steps after exposure to the sample.
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