Combinatorial Chemistry & High Throughput Screening - Volume 6, Issue 3, 2003

Fluorescence polarization was first observed in 1920 and during the next few decades the theoretical foundations of the phenomenon were clearly established. In the last two decades of the 20th century, fluorescence polarization became one of the most prevalent methods used in clinical and biomedical sciences. In this article we review the history of fluorescence polarization, its theoretical foundations and some of the more i Read More

A general survey of the analytical application of kinetic methodology in fluorescence polarization immunoassay (FPIA) is presented. Stopped-flow mixing technique (SF) allows the initial rate of the immunochemical reaction between the tracer and the antibody to be obtained, which is used as the analytical parameter instead of the equilibrium signal used in conventional FPIA. The instrumentation required is described and the f Read More

Fluorescence polarization technology has been used in basic research and commercial diagnostic assays for many decades, but has begun to be widely used in drug discovery only in the past six years. Originally, FP assays for drug discovery were developed for single-tube analytical instruments, but the technology was rapidly converted to high-throughput screening assays when commercial plate readers with equival Read More

Fluorescence polarization (FP) has become widely employed for high throughput screening used in pharmaceutical drug discovery. Assays of important signal transduction targets are now adapted to FP. In this review we examine assays for cyclic adenosine monophosphate, phosphodiesterases, and protein kinases and phosphatases using FP competitive immunoassays and a direct enzymatic method called IMAP.

This article describes some recent developments in the field of fluorescence polarization (FP) as applied to enzyme assays and single nucleotide polymorphism (SNP) genotyping. First, we present our recent progress on the application of fluorescence polarization to high throughput screening (HTS). We show how the use of a 2-thiopyridinone-based, mixed disulfide biotinylation reagent can shorten the assay time of our recentl Read More

Single nucleotide polymorphisms (SNPs) are the most abundant variations in the human genome and have become the primary markers for genetic studies for mapping and identifying susceptible genes for complex diseases. Methods that genotype SNPs quickly and economically are of high values for these studies because they require a large amount of genotyping. Fluorescence polarization (FP) is a robust technique th Read More

The effects of NaCl concentration and temperature on the rate of hybridization of complementary single-stranded DNA (24-mers) were investigated. The single label of fluorescein was used for the probe DNA. The time courses of fluorescence polarization for the probe DNA were monitored. It was shown that detection of a specific DNA sequence (24-mer) was possible in less than 10 min using fluorescence polariz Read More

Fluorescence Polarization Assays (FPAs) have been shown to have great utility in the detection of infectious diseases. Examples are presented of the use of O-polysaccharides (OPSs) for the detection of antibodies in serum, whole milk and whole blood to gram negative organisms (Brucella spp., Salmonella spp.). The use of proteins and peptides are also described for the detection of Mycobacterium bovis and Equine Infectio Read More

Antibodies raised against a given metal ion complex of a polyaminopolycarboxylate chelating agent can display specificity for the immunizing chelate and, when used in conjunction with a fluorophorelabeled analog of that chelate, can form the basis for highly sensitive and specific methods for detecting that metal ion by competitive inhibition fluorescence polarization immunoassay (FPIA). Chelate complexes of ethy Read More

Fluorescence polarization immunoassay methods for the detection of pesticides and their metabolites or degradation products are reviewed. Advantages and limitations for application to pesticide detection in environmental and food samples are discussed. The influence of the structure of fluorescentlabeled tracers and the affinity and specificity of antibodies on analytical performance is examined. The methods are simple, Read More

Successful use of fluorescence polarization assays (FPAs) in human clinical, infectious disease, and drug discovery fields has prompted us to extend its use to the grain mycotoxin field. An antibody specific to a mycotoxin and a mycotoxin-fluorophore conjugate are developed. Free toxin (extracted from the grains with a suitable solvent) competes with the toxin-fluorophore conjugate for the antibody and a change in FP re Read More