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This bead kit can be used for many applications, including:
In a typical AlphaScreen assay, one biomolecule of interested is biotinylated and associates with the Streptavidin-coated Donor beads. The other biomolecule is GST-tagged, and associates with the AlphaScreen anti-GST Acceptor beads. If the two biomolecules bind to each other, the Donor and Acceptor beads are brought into proximity. Excitation of the Donor beads causes the release of singlet oxygen, which diffuses and triggers the emission of light from the Acceptor beads when in proximity. The amount of light is directly proportional to the degree of interaction. Competition formats are also possible, using the biotinylated GST probe provided in the kit.
AlphaScreen® and AlphaLISA® are bead-based assay technologies used to study biomolecular interactions in a microplate format. The acronym ""Alpha"" stands for amplified luminescent proximity homogeneous assay. As the name implies, some of the key features of these technologies are that they are non-radioactive, homogeneous proximity assays. Binding of molecules captured on the beads leads to an energy transfer from one bead to the other, ultimately producing a luminescent/fluorescent signal. To understand how a signal is produced, one must begin with an understanding of the beads. AlphaScreen and AlphaLISA assays require two bead types: Donor beads and Acceptor beads. Each bead type contains a different proprietary mixture of chemicals, which are key elements of the AlphaScreen technology. Donor beads contain a photosensitizer, phthalocyanine, which converts ambient oxygen to an excited and reactive form of O2, singlet oxygen, upon illumination at 680 nm. Please note that singlet oxygen is not a radical; it is molecular oxygen with a single excited electron. Like other excited molecules, singlet oxygen has a limited lifetime prior to falling back to ground state. Within its 4 µsec half-life, singlet oxygen can diffuse approximately 200 nm in solution. If an Acceptor bead is within that proximity, energy is transferred from the singlet oxygen to thioxene derivatives within the Acceptor bead, subsequently culminating in light production at 520-620 nm (AlphaScreen) or at 615 nm (AlphaLISA). In the absence of an Acceptor bead, singlet oxygen falls to ground state and no signal is produced. This proximity-dependent chemical energy transfer is the basis for AlphaScreen's homogeneous nature.
|Product Brand Name||AlphaScreen|
|Shipping Condition||Blue Ice|
|Unit Size||500 assay points|
Gß? is biotinylated and binds to streptavidin-donor beads, while cGIRK1 binds to anti GST-acceptor beads via its GST tag. Specific interaction between the two proteins leads to signal increase. The binding is fully reversible and completely abolished by competition with G protein a subunit. This assay can be easily applied to any protein-protein interaction.
Alpha has been used to study a wide variety of interactions, including protein:protein, protein:peptide, protein:DNA, protein:RNA, protein:carbohydrate, protein:small molecule, receptor:ligand, and nuclear receptor:ligand interactions. Both cell-based and biochemical interactions have been monitored, and applications such as phage display, ELISA, and EMSA (electrophoretic mobility shift assay) have been adapted to Alpha.
AlphaScreen® and AlphaLISA® are bead-based assay technologies used to study biomolecular interactions in a microplate format. The acronym “Alpha” stands for Amplified Luminescent Proximity Homogeneous Assay. The assay does not require any washing steps. Binding of proteins or other binding partners captured on the beads leads to an energy transfer from one bead to the other, ultimately producing a luminescent signal.