Fluorescence-activating and absorption-shifting tag
FAST, which stands for Fluorescence-Activating and absorption-Shifting Tag, is a small, genetically encoded protein tag that reports where a protein of interest is inside cells. It is about 14 kDa (125 amino acids) and was engineered from the photosensitive PYP protein. FAST itself does not glow; it binds a fluorogenic dye that is nonfluorescent until it attaches to FAST. When bound, the pair becomes bright and changes how they absorb light, giving a clear, targeted signal with high labeling specificity.
FAST was first described in 2016 by researchers at the Ecole normale supérieure in Paris. Since then, it has evolved into useful derivatives, including splitFAST (2019), a system that reports protein–protein interactions by splitting FAST into two parts that light up only when the tagged proteins interact. In 2023, CATCHFIRE appeared, a self-reporting, reversible dimerizing system that fuses FAST-based domains to two proteins of interest and uses fluorogenic inducers to drive their proximity, boosting fluorescence dramatically.
In the broader context, fluorescence imaging is widespread in biology, traditionally dominated by GFP-like proteins. FAST and related chemical-genetic approaches offer an alternative: a protein tag gives precise labeling and localization, while an external dye supplies fluorescence that is activated only upon binding. A range of chromophores can be used, from natural ones like FMN, biliverdin, and bilirubin to synthetic dyes compatible with tags such as SNAP-, CLIP-, and HaloTag. The FAST system uses a fluorogenic chromophore derived from 4-hydroxybenzylidene rhodanine; it is nonfluorescent on its own, but becomes bright and shifts its absorption when bound to FAST.
Many FAST variants exist, including Y-FAST, iFAST, pFAST, greenFAST, redFAST, frFAST, nirFAST, nanoFAST, and combinations in FAST dimers. Different fluorogens offer different emission colors, brightness, and binding affinity, and some are nonpermeant, allowing selective labeling of membrane or extracellular proteins to study trafficking from synthesis to excretion. FAST is advancing toward near-infrared reporting, which is ideal for imaging whole organisms due to deeper tissue penetration, reduced photodamage, and higher signal-to-noise.
The FAST sequences are available on FPbase. splitFAST provides a reversible, fluorescent readout of transient protein–protein interactions in living cells by using two fragments, NFAST (about 114 amino acids) and CFAST (10–11 amino acids). When their target proteins interact, the two pieces reassemble FAST, bind a fluorogen, and illuminate the interaction. This approach serves as an accessible alternative to FRET or BiFC for studying dynamic associations and disassembly.
CATCHFIRE extends this concept by fusing proteins of interest to FAST-based dimerizing domains, FIREtag and FIREmate. Fluorogenic inducers (the “match” series) trigger their interaction and dramatically boost fluorescence. The system is reversible, enabling real-time observation of induced interactions and making it possible to explore protein localization, trafficking, organelle transport, and other cellular processes with high spatial and temporal resolution. CATCHFIRE also points to a new class of fluorogenic biosensors for signaling and apoptosis studies.
FAST and its derivatives are used across a wide range of applications: monitoring protein trafficking and interactions, designing biosensors, and enabling chemically induced dimerization. They work in fluorescence microscopy, flow cytometry, and other fluorescence-based methods, including super-resolution imaging of living cells. FAST is particularly valuable in low-oxygen environments, where many other fluorescent proteins struggle, and has been applied to anaerobic microbes, certain archaea, and a variety of pathogens. It’s also used in fungi and mammalian cells to study cellular mechanisms such as receptor signaling, antiviral proteins, and membrane contact sites between organelles.
Overall, FAST and splitFAST provide flexible, reversible, and highly selective tools for studying the dynamic behavior of proteins in living systems, with ongoing developments expanding their color range, brightness, and range of biological applications.
This page was last edited on 3 February 2026, at 04:20 (CET).