|Assay Principle of the AlphaScreen® SureFire® Phospho-ERK1/2 assay|
Amplified Luminescent Proximity Homogeneous Assay (Alpha) technology represents a distinct approach to sensing proximity; which can be used to study protein-protein interactions. Unlike FRET or BRET, which use resonant energy transferred from a fluorescent or bioluminescent donor respectively, AlphaScreen® uses diffusion of singlet oxygen from donor to acceptor beads. When donor beads are excited at a wavelength of 680 nm; oxygen is converted to singlet oxygen. If the acceptor bead is in proximity; the singlet oxygen initiates a series of chemical reactions that produce a luminescent signal with a wavelength in the range of 520-620 nm. Since singlet oxygen has a short lifetime the donor and acceptor must be within 200 nm of one another. This technology has been employed in AlphaScreen® and AlphaLisa® applications that are based on the sandwich ELISA approach.
The AlphaScreen® laser that BMG LABTECH employs for excitation was first developed in the PHERAstar. Using a laser is helpful as an intense light source may be required to generate a signal. The same laser has now been implemented in the CLARIOstar. This should open the door to many life science labs to be able to work with this powerful technology.