AlphaLISA immunoassays are homogenous bead-based luminescent amplification proximity assays that are highly sensitive and offer a wide dynamic range while requiring only small sample sizes. This proximity assay generates a luminescent or fluorescent signal after energy transfer occurs between molecule-bound Acceptor and Donor beads. Donor beads contain the photosensitizer phthalocyanine, which converts ambient oxygen into a reactive singlet oxygen species when illuminated at 680 nm. The excited singlet oxygen has a half-life of 4 µsec and can diffuse approximately 200 nm in solution. If an Acceptor bead is within this range, the energy is transferred and converted into a measurable luminescent signal at 615 nm. In the absence of Acceptor beads, the singlet oxygen falls to its ground state and no signal is produced.

These assays are versatile and compatible with a variety of sample types, including serum, plasma, cerebrospinal fluid (CSF), tissue extracts, cell culture supernatants, and cell lysates. They can be applied to diverse research areas such as enzyme activity studies, receptor-ligand interactions, ion and second messenger signaling, as well as investigations involving DNA, RNA, proteins, protein–protein interactions, peptides, and small molecules. In addition, AlphaLISA assays are adaptable to high-throughput formats, with options for 96-, 384-, and 1536-well plates, making them ideal for large-scale screening applications.