Mass-sensing, multianalyte microarray immunoassay with imaging detection

Miniaturization of ligand binding assays may reduce costs by decreasing reagent consumption, but it is less apparent that miniaturized assays can simultaneously exceed the sensitivity of macroscopic techniques by analyte "harvesting" to exploit the total analyte mass available in a sample....

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Bibliographic Details
Published in:Clinical chemistry (Baltimore, Md.) Md.), 1998-09, Vol.44 (9), p.2036-2043
Main Authors: Silzel, John W, Cercek, Bibijana, Dodson, Charles, Tsay, Tsong, Obremski, Robert J
Format: Article
Language:English
Subjects:
Avidin
Biological and medical sciences
Biotin - chemistry
Fluorescence
Fluorescent Dyes
Fundamental and applied biological sciences. Psychology
Fundamental immunology
Humans
Image Enhancement
Immunoassay - methods
Immunoglobulin G - analysis
Miniaturization
Molecular immunology
Sensitivity and Specificity
Techniques
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Summary:Miniaturization of ligand binding assays may reduce costs by decreasing reagent consumption, but it is less apparent that miniaturized assays can simultaneously exceed the sensitivity of macroscopic techniques by analyte "harvesting" to exploit the total analyte mass available in a sample. Capture reagents (avidin or antibodies) immobilized in 200-microm diameter zones are shown to substantially deplete analyte from a liquid sample during a 1-3-h incubation, and the assays that result sense the total analyte mass in a sample rather than its concentration. Detection of as few as 10(5) molecules of analyte per zone is possible by fluorescence imaging in situ on the solid phase using a near-infrared dye label. Single and multianalyte mass-sensing sandwich array assays of the IgG subclasses show the sensitivity and specificity of ELISA methods but use less than 1/100 the capture antibody required by the 96-well plate format.
ISSN:0009-9147
1530-8561
DOI:10.1093/clinchem/44.9.2036