AlphaScreen (Amplified Luminescent Proximity Homogeneous Assay) is a very sensitive, non-radioactive homogeneous assay technology. It has successfully been used for enzyme assays (kinase, helicase, protease, ), protein interaction assays (ligand/receptor, protein/protein, protein/DNA), immunoassays, and GPCR functional assays (cAMP, IP3).
AlphaScreen relies on the use of "Donor" and "Acceptor" beads that are coated with a layer of hydrogel providing functional groups for bioconjugation. When a biological interaction between molecules brings the beads into proximity, a cascade of chemical reactions is initiated to produce a greatly amplified signal. Upon laser excitation, a photosensitizer in the "Donor" bead converts ambient oxygen to a more excited singlet state. The singlet state oxygen molecules diffuse across to react with a chemiluminescer in the Acceptor bead that further activates fluorophores contained within the same bead. The fluorophores subsequently emit light at 520-620 nm.
In the absence of a specific biological interaction, the singlet state oxygen molecules produced by the Donor bead go undetected without the close proximity of the Acceptor bead.
PerkinElmer can label your target protein or ligand (binding partner) with biotin, fluorescein or digoxigenin and cross-link it to AlphaScreen Donor beads. AlphaScreen Acceptor beads are labeled with your purified protein of interest. These enable you to create your own tailored AlphaScreen assay.
Labeling of Target Protein/Ligand
The target protein or ligand is labeled with biotin, fluorescein or digoxigenin. A minimum of 0.25 mg of purified protein (concentration > 1 mg/mL) in PBS is required. The level of label per target protein is determined using protocol developed in-house, and activity of the target protein is verified using a customer-provided protocol and reagents. The purified labeled protein is sent back to you within 1-2 weeks with a Q/C data sheet.
Acceptor Bead Labeling
The target protein is cross-linked to the AlphaScreen Acceptor beads. A minimum of 0.25 mg of purified protein (concentration > 1 mg/mL) in PBS is required. The level of cross-linked protein is determined using an appropriate assay, and the activity of the cross-linked protein is determined using customer-provided protocol and reagents. The purified modified Acceptor beads are sent back to you within 1-2 weeks with a Q/C data sheet.