Fluorescence immunoassay is a sensitive technique that can be used in the measurement of many compounds, including drugs, hormones, and proteins; in the identification of antibodies; and in the quantification of antigens such as viral particles and, potentially, bacteria. Homogeneous fluorescence immunoassay, fluorescent excitation transfer immunoassay, fluorescence polarization immunoassay, solid-phase "dipstick" immunoassay, solid-phase microbead fluorescence immunoassay, substrate-labeled fluorescence immunoassay, and fluorescence immunoassays using internal reflectance spectroscopy or phycobiliprotein conjugates are reviewed.
"Specific DNA sequences have been detected by hybridization assays using different materials such as biotin-avidin, protein-enzyme, and fluorescent dyes. However, these methods are limited by low signal intensity, rapid photobleaching as well as biosafety problems [6,7]. Fluorescent semiconductor nanocrystals, quantum dots (QDs) possess unique properties and have significant advantages (such as tunable band gap and extraordinary photostability) over the classic organic dyes. "
[Show abstract][Hide abstract] ABSTRACT: In the present study we describe sandwich design hybridization probes consisting of magnetic particles (MP) and quantum dots (QD) with target DNA, and their application in the detection of avian influenza virus (H5N1) sequences. Hybridization of 25-, 40-, and 100-mer target DNA with both probes was analyzed and quantified by flow cytometry and fluorescence microscopy on the scale of single particles. The following steps were used in the assay: (i) target selection by MP probes and (ii) target detection by QD probes. Hybridization efficiency between MP conjugated probes and target DNA hybrids was controlled by a fluorescent dye specific for nucleic acids. Fluorescence was detected by flow cytometry to distinguish differences in oligo sequences as short as 25-mer capturing in target DNA and by gel-electrophoresis in the case of QD probes. This report shows that effective manipulation and control of micro- and nanoparticles in hybridization assays is possible.
[Show abstract][Hide abstract] ABSTRACT: An immunoassay format is presented that takes advantage of the microfluidic properties of the H-FilterTM for measuring sample analyte concentration. The method relies on the diffusion of analyte particles into a region containing beads coated with specific antibody. Competitive binding of labeled analyte and sample analyte with a limited number of binding sites allows measurement of the concentration of sample analyte based on the fraction of labeled analyte bound. The fraction of labeled analyte bound can be determined with a microcytometer by measuring the bead fluorescence intensity on the microcytometer portion of an integrated microfluidic chip. It is not necessary to separate the beads from the mixture because the bead intensity can be determined above the background of unbound labeled antigens. Other advantages include the ability to eliminate large interfering particles from samples, continuous sample monitoring, and the ability to concentrate the beads. Microfluidic immunoassay formats also consume smaller volumes of costly reagents and sample.
Proceedings of SPIE - The International Society for Optical Engineering 08/1999; DOI:10.1117/12.359334 · 0.20 Impact Factor
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