Multi-analyte analysis of biological fluids with a recycling immunoaffinity column array.
A system for isolating and measuring up to 30 analytes in a single biological sample is described. The system is based on recycling a pre-labeled sample through an array of capillary immunoaffinity columns, each packed with glass beads, coated with a different antibody, thus enabling each column to isolate and extract a single analyte. Detection of the bound analytes is achieved by laser-induced fluorescence (LIF), using a laboratory-built scanning detector coupled to a fiber-optic spectrometer. The array can be regenerated up to 200 times, provided a suitable temperature is maintained. The individual immunoaffinity columns are able to bind between 2.9 and 3.6 ng of analyte, depending upon the individual column, with lower limits of detection (LOD) in the order of 1.6-2.8 pg/ml. The inter- and intra-assay coefficients of variation (CV) for all 30 columns in the array were less than 6.03+/-0.33 at analyte concentrations of 100 pg/ml. Comparison to standard enzyme-immunoassays demonstrated r(2) values in the range of 0.9151-0.9855 when analyzed by least-squares linear regression.
Phillips
T M
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Ultramicro Analytical Immunochemistry Resource, Division of Bioengineering and Physical Sciences, Office of Research Services, OD, National Institutes of Health, Bldg 13/3E42, 9000 Rockville Pike, Bethesda, MD 20892, USA. phillipt@ors.od.nih.gov
D000906:Antibodies / Q000302:isolation & purification; D001826:Body Fluids / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods*; D017743:Equipment Reuse; D006801:Humans; D007124:Immunoenzyme Techniques; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000379:methods; D013053:Spectrophotometry / Q000379:methods
D000906:Antibodies