| Acceptor |
In resonant energy transfer, the molecule that receives the energy transfer and emits a photon. |
| Background |
Raw signal when assay activity is zero. |
| Channel A |
Fluorescence measured through the filter chosen to measure acceptor fluorescence; increases when FRET occurs. |
| Channel B |
Fluorescence measured through the filter chosen to measure donor fluorescence; decreases when FRET occurs. |
| Donor |
In resonant energy transfer, the molecule that absorbs a photon and initiates energy transfer to the acceptor. |
| Fluorescence |
Energy emitted as photons when excited molecules return to their ground state. |
| Fluorophore (chromophore) |
The chemical moiety of a dye molecule which absorbs light and, in the case of a fluorophore, emits light through fluorescence. |
| FRET |
Fluorescence Resonance Energy Transfer- In two adjacent dyes, the energy of an excited electron in one dye (the donor) is passed to an electron of an adjacent dye (the acceptor) through resonance, then released as a photon. |
| HTRF® |
Homogeneous Time-Resolved Fluorescence, a trademark of Packard Instruments. |
| Noise |
Statistical variability in the response of the assay from all sources, including: variation in actual quantities of reagents added or their concentration, instrument output variability, geometry of plates, uncertainty or variability in background, etc. |
| Phycobiliproteins |
Photosynthetic accessory pigments found in cyanobacteria, red algae, and cryptomonads that absorb light in spectral regions where chlorophyll absorbs poorly and transfers energy to chlorophyll for use in photosynthesis. Chromophores (linear tetrapyrroles called bilins) are attached to a protein framework. |
| Proximity pair |
Two fluorescent dyes selected for their ability to successfully achieve FRET when brought into close proximity in a FRET Assay or in a tandem dye. |
| Quantum efficiency |
Proportion of absorbed photons released as fluorescence. |
| Quench |
absorption of light in an assay mixture by either reactants or impurities. |
| Raw signal |
Signal before correction for background. |
| Self-quenching |
Tendency of fluorophores with small Stokes shifts to absorb photons at some of the same wavelengths at which they emit. |
| Signal |
The "real" response of an assay to activity in the sample, independent of all sources of error and statistical variability, corrected for any background. |
| Stokes' shift |
Difference in wavelength between the fluorescence emission and excitation peaks of a given dye. |
Tandem fluorescent dye
(tandem dye, tandem conjugate) |
Two fluorescent dyes capable of FRET covalently coupled. Tandem dyes are used when increased Stokes shifts or multiple emission wavelengths from a single excitation wavelength are desired. |
