Peer-reviewed and preprint literature applying Twinkle fluorogenic reporters (FAST, splitFAST, CATCHFIRE and variants). Search or filter by reporter, type, and year; each entry links to the publisher via DOI.
A Synthetic Promoter Library Spanning a Wide Dynamic Range for Metabolic Pathway Engineering in Zymomonas mobilis
A synthetic promoter library for Zymomonas mobilis, 29 promoters spanning about 500-fold. FAST, written Y-FAST here and read with TFLime, is the reporter they switch to for the anaerobic screen because sfGFP needs oxygen, and the promoter ranking survives the switch with an R2 of 0.95 against sfGFP. First reported use of FAST in this organism.
Imaging Membrane Contact Sites Dynamics: Matching Reporter Properties to Biological Questions
A reporter-by-reporter comparison of contact site imaging, written by the SPLICS group, so by the authors of the main competing technology. split-FAST is one of four named classes and comes out of their comparative table with the shortest limitation list, a single entry, the need to add a fluorogen, against slow maturation and poor reversibility for split-GFP. Their conclusion is a division of labour rather than a ranking: fast reversible reporters such as split-FAST for resolving formation, dissociation and turnover in live cells, split-GFP for long-lived abundance and spatial organisation. They rate split-FAST in vivo applicability only moderate, which is a fair criticism. 139 references, with association and dissociation kinetics and dissociation constants given per class.
Tri-Organelle Contact Sites Integrate Metabolic Function in Health and Disease
A review of Mito-ER-lipid droplet tri-organelle contact sites. Its section 7.3 frames contact site biosensors in three generations and states that irreversible split-GFP permanently tethers the two organelles, while the low-affinity splitFAST variant used in FABCON detects contacts with minimal perturbation, confirmed by electron microscopy. Names splitFAST-based FABCON and PRINCESS as the premier tools for monitoring physiological transitions without structural artifact. No Twinkle author, no Gautier citation: independent third-party support for the non-perturbing split reporter argument.
Non-Covalent Reversibly Photoconvertible Fluorescent Tags for Wash-Free Protein Labeling
Showed that FAST-family tags turn into negative reversible photoswitchers when fluorogen concentration is lowered and light intensity raised, and that pairing two fluorogens yields positive photoswitchers and non-covalent reversibly photoconvertible tags that change color under light. Exploiting this light-driven color change in two spectral channels, the authors distinguish up to six spectrally similar FAST variants in live cells.
Reversible, Chemically Gated FRET via Ligand-Activated Acceptors
First crystal structures of apo and ligand-bound FAST, showing that N-terminal residues occlude the empty pocket and are displaced on fluorogen binding to form a water-stabilized cavity. Different hydroxybenzylidene rhodanine fluorogens tune the binding geometry and yield single or multiexponential fluorescence lifetimes, which the authors exploit to use FAST as a chemically gated, reversible FRET acceptor for ligand-controlled FLIM-FRET, mapping MAGUK scaffold organization in live cells.
Unexpected functional role of the transactivation domain for nuclear import of STAT5
Used CATCHFIRE to reveal an unexpected role of the STAT5 transactivation domain in nuclear import.
A sensitive interactome-based approach to characterize drug activity in living human cells
splitFAST chosen for what BiFC cannot do. The authors note that bimolecular fluorescence complementation is irreversible and therefore cannot report a drug destabilising a complex that has already formed, so they introduce splitFAST2 for that half of the work and read drug-sensitive ERK1 interactions in living cells. Reversibility is the stated reason for the choice.
High-throughput screen identifies a potent MCU activator boosting cardiac contractile bioenergetics
Drug discovery reaches the contact site. A high-throughput screen identifies CGP7930 as an activator of the mitochondrial calcium uniporter, binding MICU1 at a pocket between its N and C lobes, and the compound then boosts calcium-dependent mitochondrial energy metabolism in cardiomyocytes and contractile performance in mouse hearts. The contact-site measurement is a single clean experiment: HeLa cells expressing ER-Mit RspA-splitFAST with the Lime fluorogen at 4 micromolar, where one hour of CGP7930 significantly raised the proportion of mitochondria in contact with the ER, which the authors read as a second mechanism for the drug beyond the uniporter itself. Two things to note for us: it is the second paper in the corpus to name the Lime fluorogen, and Paloma Garcia-Casas, first author of PRINCESS, is among the authors, so the reporter travelled with its builder rather than through a catalogue. Preprint, not peer reviewed.
Evaluating the far-red emitting fluorescence-activating and absorption-shifting tag (frFAST) chemogenetic labeling system for live-cell super-resolution microscopy
Reported far-red chemogenetic FAST labeling approaches for background-free super-resolution imaging of intracellular targets.
Excessive EFHD1-dependent ER-mitochondrial contacts drive a maladaptive antiviral response in metabolic liver disease
Used splitFAST revealed by TF-Lime to count ER-mitochondria contacts and time their lifetime live in HepG2 cells, showing that the Ca2+-dependent actin cross-linker EFHD1 stabilizes these contacts. In metabolic liver disease (MASH), EFHD1 excess makes contacts persist, fragments mitochondria and triggers a maladaptive antiviral stress response; inhibiting EFHD1 protects hepatocytes.
Small molecules for genetically encoded control and imaging of protein proximity
Review of chemically-induced proximity (CIP): small-molecule inducers that conditionally recruit proteins, tracing their evolution from non-covalent to covalent and hybrid systems, and the recent shift to single scaffolds that integrate proximity induction with optical reporting, highlighting CATCHFIRE as one of the tools most closely uniting the two.
Kinetic and Mechanistic Characterization of RSpFAST, a Non-Covalent Versatile Chemogenetic Platform for Fluorescence Imaging
Introduces RSpFAST (a pFAST tag paired with a reversibly photoisomerizable fluorogen) and characterizes its thermokinetics, photochemistry and structure. Reveals a dual regime: a stable wash-free label becomes a negative photoswitcher at low fluorogen concentration and high light (photoejection-driven), enabling dynamic-contrast and super-resolution imaging in live and fixed cells.
HIV-1 Nef Homodimerization as a Structural Mechanism for Kinase Activation and Small Molecule Inhibitor Action
Showed that mutating the HIV-1 Nef dimer interface residues Leu112 and Phe121 abolishes dimerization while preserving the Nef fold, giving monomeric complexes with Hck kinase domains. Used a splitFAST complementation assay to provide direct evidence that small-molecule inhibitors disrupt Nef homodimer formation in solution.
Lighting Up Biology: GFP-Inspired Fluorescent Probes for Sensing and Imaging
A perspective on probes built from the GFP chromophore core, and the richest of the grade-2 reviews on us: thirteen sentences covering the HBR activation mechanism, the 14 kDa size, specificity and reversibility as properties, pFAST credited to the Gautier group, splitFAST's purpose, and a figure panel on FAST's own absorption and emission spectra.
Advanced imaging of membrane contact sites
A 151-reference methods review of membrane contact site imaging, from electron microscopy and super-resolution to FRET, split-GFP/SPLICS and splitFAST. It treats reversible splitFAST reporters and PRINCESS as the way to read contact dynamics together with the Ca2+ signals they carry, and names the missing tool explicitly: no reporter yet tracks lipid transfer at contacts in real time, which the authors call a priority.
Discerning protein pools by selective staining with self-labeling tags
Review of how self-labeling tags separate protein populations: surface, intracellular and endocytosed pools of transmembrane proteins, using permeable versus impermeable dye-ligands and cleavable groups that strip the remaining surface pool. Covers SNAP-tag, HaloTag and FAST. For FAST, sulfonated hydroxybenzylidene rhodanine fluorogens are impermeable and were used to follow trafficking of the dopamine D2 receptor by flow cytometry.
Intrafilament nucleotide exchange in a prokaryotic actin homolog
The bacterial cytoskeleton enters the corpus, and pFAST is the label that made it visible. MreB, the actin homolog that sets bacterial cell shape, is fused to pFAST and watched as single double filaments assemble on a lipid monolayer, by total internal reflection microscopy with the TFLime ligand, beside atomic force and electron microscopy. The finding is that nucleotide exchange happens inside an assembled filament, which actin does not do, its subunits keeping their nucleotide trapped until they depolymerise. Two points for us beyond the biology. The authors state that the pFAST-MreB fusion is fully functional in vivo, which is the kind of sentence a tag vendor wants in print. And their methods cite our catalogue numbers outright, 5 micromolar of either TFLime (P-480541-250, The Twinkle Factory) or TFCoral (516600250, The Twinkle Factory), which makes this the clearest customer evidence the corpus holds. Preprint, not peer reviewed.
Chemically Responsive Protein Switches for the Precise Control of Biological Activities
Used CATCHFIRE chemically-induced dimerization to build reversible protein switches: assembling split enzymes (luciferases, proteases, DNA recombinases) and a titratable inducible gene-expression system (CATCH-ON, via GAL4/p65Δ) to control activities including insulin secretion and suicide switches.
A characterized promoter suite enables metabolic rewiring of the acetogen Acetobacterium woodii toward ethanol production
Developed a modular plasmid platform and a suite of characterized native, heterologous and methanol-inducible promoters for the acetogen Acetobacterium woodii, using pFAST with HMBR as the anaerobe-compatible reporter to quantify promoter strength. Replacing the native adhE promoter and deleting the neighbouring LysR-type regulator shifted the strain from acetogenic toward partly ethanologenic, yielding ethanol from fructose and from the C1 substrates methanol plus CO2 and formate plus syngas.
Chemogenetic timestamping for the precise tracing of cell history into protein assemblies
A preprint on chemogenetic 'timestamping' to trace cell history into protein assemblies.
Advancing the monitoring of organelle contact sites in vitro and in vivo
A second review from the SPLICS group, Barazzuol, Peggion, Brini, Nasu and Calì, on monitoring organelle contact sites in vitro and in vivo. Same standing as Poggio: the authors of the competing split-GFP contact reporters assessing the field, with split-FAST and the full tag name both named.
Chemigenetic fluorescent biosensors in biological imaging: new trends and advances
Review of chemigenetic fluorescent biosensors, covering how the two most widely used systems, HaloTag and FAST, have been built into biosensors. Highlights bright, photostable far-red fluorophores for in vivo and multiplexed imaging, fluorescence lifetime readouts that make imaging quantitative, and chemigenetic integrators that record physiological events for later analysis.
The fluorescence‐activating and absorption‐shifting tag (FAST), a versatile protein marker for live plant cell imaging
Established FAST, greenFAST/redFAST and splitFAST for versatile multicolor imaging in plant cells.
A coordinated transcriptional program controls de novo Golgi biogenesis
CATCHFIRE used inside a mechanistic cell-biology study rather than a methods paper: the authors ablate the Golgi, watch it rebuild, and run a CATCHFIRE-based retrograde Golgi-to-ER trafficking assay with FIREMate and FIREtag to show that retrograde transport from newly forming Golgi is slowed. A dedicated methods section describes the assay.
Establishing the fluorescence-activating and absorption-shifting tag as a fluorescent reporter
A preprint establishing FAST as a fluorescent reporter in a new host system.
Genetically Encoded Tools to Monitor and Interrogate Membrane Contact Sites
A review of the genetically encoded reporters available for membrane contact sites, split-FAST among them with thirteen mentions, set beside split-GFP and the dimerization-dependent fluorescent proteins. Published in Contact, like Mukunda, and completes the 2026 set of contact-site reporter reviews alongside Filadi and Poggio.
Chemogenetic Modulation of Luciferase Emission Color for Imaging and Sensing
Introduced LumiFAST, a chemogenetic way to tune luciferase emission color, for bioluminescence imaging and sensing.
Translational antagonism enables bet-hedging in yeast via mitochondrial bistability
Used splitFAST to report cytochrome c oxidase assembly in single yeast cells, showing that a bistable switch in mitochondrial activity splits a population into fermenting arrestors and respiring recoverers.
Enabling Plasmid-Based Expression in Clostridium kluyveri Using a Biparental Methylation-Conjugation System
Built a biparental methylation-conjugation system that bypasses restriction-modification barriers in Clostridium kluyveri DSM 555T, enabling stable plasmid delivery, and used FAST with HMBR as an oxygen-independent reporter to confirm heterologous expression (peak emission 542 nm, measured by plate reader). Opens C. kluyveri, a chain-elongating biocatalyst for medium-chain fatty acids, to metabolic engineering.
Monitoring intracellular antibiotic concentrations in real-time using allosteric biosensors
Used a FAST-based allosteric biosensor to monitor intracellular antibiotic concentrations in real time.
Development of carbon monoxide-tolerant fluorescent reporter system functional under anoxic conditions in Eubacterium callanderi KIST612
Built a fluorescent reporter system for the acetogen Eubacterium callanderi KIST612 that works under strict anoxia and tolerates carbon monoxide, after characterizing the strain autofluorescence and selecting FAST with HMBR as the best-suited reporter. FAST-expressing strains were about 4.5 times brighter than controls in both glucose and CO conditions, and a CO-sensing construct gave fluorescence proportional to CO partial pressure.
Random Mutagenesis-Based Optimization of the Fluorogen-Activating Protein picoFAST
Optimized picoFAST, an ultra-small fluorogen-activating protein, by random mutagenesis.
Inactive ryanodine receptors sustain lysosomal availability for autophagy by promoting ER-lysosomal contact site formation
Used splitFAST to study how inactive ryanodine receptors sustain lysosomal autophagy in Alzheimer's models.
Découverte du rôle des mitochondries dans la différenciation des cellules en neurones
A Le Monde news article on the discovery of mitochondria's role in neuron differentiation (Twinkle-team work).
Reversible Chemogenetic Fluorescence Labeling with pFAST in C. elegans
Made a tandem pFAST (td-pFAST) C. elegans strain for reversible multicolor chemogenetic labeling in worms.
Bathochromic Shift via C=O to C=S Substitution: A Far-Red Fluorogen for Multiplexed FLIM with FAST Fluorogen-Activating Protein
Extended the FAST palette into the far-red with HBTR-3,5-DOM, a fluorogen made by C=O-to-C=S substitution.
Imaging and quantifying organelle contact sites using a reversible splitFAST complementation assay
A methods chapter on imaging and quantifying organelle contact sites with a reversible splitFAST complementation assay.
Anaerobic Fluorescent Reporters for Studying Pseudomonas aeruginosa-Specific Responses in a CF Polymicrobial Assay
A PhD thesis developing anaerobic FAST reporters to study Pseudomonas aeruginosa responses.
Low-affinity exchangeable probes for super-resolution and long-term fluorescence imaging
An outside landscape review that puts FAST in one table with DNA-PAINT, RNA aptamers, exchangeable HaloTag ligands (xHTL), RhoTag, SiR-Tag, the de novo NovoTags and the scFv FAPs, and judges them on eight criteria. It is the clearest third-party statement we hold that fast exchange is an advantage rather than a defect: unlimited labelling, minimal photobleaching and ON/OFF switching for super-resolution. It credits FAST with a 14 kDa reversible tag, 28 nm SMLM on microtubules in its benchmark table, colour switching by fluorogen exchange, and a quantitative signal held for hours in low-oxygen biofilms where GFP-like proteins fail for want of oxygen. It is equally plain about the limits: protein tags and aptamers are rated moderate in super-resolution next to DNA-PAINT, the suggested cause being photodamage of the binding site capping the number of binding and unbinding cycles, with slow dye exchange, poor atom economy and the need for transfection as the other costs.
Unequal mitochondrial segregation promotes asymmetric fates during neurogenesis
Used live imaging with Twinkle-team tools to show unequal mitochondrial segregation biases neuron-versus-progenitor fate in neurogenesis.
Early HIV-1 maturation drives Env clustering and fusion competence
Studied how early HIV-1 maturation drives Env clustering and fusion competence, using FAST-family imaging.
GLP-1R associates with VAPB and SPHKAP at ERMCSs to regulate β-cell mitochondrial remodelling and function
The GLP-1 receptor axis enters the corpus. Working in rat INS-1 832/3 beta cells, the authors show that the GLP-1 receptor associates with VAPB and SPHKAP at ER-mitochondria contact sites, and that stimulation with exendin-4 remodels mitochondria and reduces the number of contacts per mitochondrial area. The contact count is read with splitFAST2, using the PRINCESS constructs ER-RspA-NFAST and OMM-short-RspA-CFAST, in wild-type INS-1 832/3 cells only, vehicle against exendin-4 at 1 micromolar for 10 minutes, 50 to 57 cells over three independent experiments. The GLP-1R knockout and SPHKAP knockdown arms belong to the electron microscopy of contact length, not to the splitFAST count: the two readouts are in the same figure and are easy to merge by mistake. Two things make it unusual for us. It is the first paper in the corpus whose methods name The Twinkle Factory as the source of the fluorogen, the Lime splitFAST ligand at 5 micromolar, while the plasmids come from the Padua group: the gene travels through the literature, the molecule is bought. And it puts a FAST reporter inside the pharmacology of GLP-1 receptor agonists, a field the corpus did not touch.
Chemigenetic Approaches for the Development of Fluorescent Biosensors for Biological Imaging
A review of chemigenetic (hybrid) fluorescent biosensors, including FAST-based designs.
Super-Resolution Live-Cell Mapping of Protein-Protein Interactions Using Chemogenetic splitFAST
Mapped protein-protein interactions at super-resolution in live cells using chemogenetic splitFAST.
A color-tailored fluorogenic sensor for hydrogen peroxide
Built HyPerFLEX, a FAST-based color-tunable fluorogenic sensor for high-precision H2O2 detection in living cells.
Thermophilic Chassis-Enabled High-Throughput Selection of a Thermostable Fluorogenic Reporter
Selected thermostable FAST variants (hsFAST/tsFAST) via a thermophilic-chassis high-throughput screen.
Human MX1 orchestrates the cytoplasmic sequestration of neo-synthesized influenza A virus vRNPs
Applied splitFAST to dissect how human MX1 sequesters newly made influenza vRNPs in the cytoplasm.
Fluorescence Lifetime-Based Separation of FAST-Labeled Cellular Compartment
A step-by-step protocol for FAST-based fluorescence-lifetime imaging (FLIM) to separate labels sharing a color channel.
Synthetic protein degradation circuits using programmable cleavage and ligation by Sortase A
Used splitFAST as the readout of a peptide bond being made or broken: Sortase A ligates the two fragments to switch fluorescence on, and cleaves a full-length FAST carrying an LPETGGG motif to switch it off, driving logic-gated protein degradation in live HeLa cells.
Type IV Pili-Associated Secretion of a Biofilm Matrix Protein From Clostridium perfringens That Forms Intermolecular Isopeptide Bonds
Characterized type IV pili-associated secretion of a biofilm matrix protein in Clostridium perfringens, using FAST.
Characterization of Fluorescent Reporters for Flow Cytometry-Based Single-Cell Studies in Saccharomyces cerevisiae
Benchmarked green/red/blue fluorescent reporters, including frFAST, for flow-cytometry single-cell studies in yeast.
CATCHFIRE-Based Versatile and High-Throughput Screening for Protein Secretion in Bacillus subtilis
Used CATCHFIRE to build a high-throughput screen for protein secretion in Bacillus subtilis.
Design and evaluation of a tripartite chemogenetic fluorescent reporter for visualizing ternary protein complexes
Engineered a tripartite (three-fragment) splitFAST that lights up only when three proteins assemble, visualizing ternary complexes in live cells.
A guide to characterizing the dynamic mitochondria-endoplasmic reticulum contact sites
A review guide to characterizing mitochondria-ER contact sites, referencing reversible splitFAST-type reporters.
Key challenges and recommendations for defining organelle membrane contact sites
The consensus paper of the contact-site field, written by twelve of its leaders including Calì and Brini at Padova, Kornmann, Hajnóczky, Rizzuto and Scorrano, and the current successor to the 2019 consensus. It defines what counts as a contact site and how one should be measured, and it treats splitFAST at length, giving the fragment boundaries, the reversibility, PRINCESS, and a list of served contacts longer than ours that includes the nucleus. Two things in it are not held anywhere else in the corpus: linker ranges beyond our 12 and 25 nm, namely linear linkers of 10 and 40 nm, and a limitation stated in the consensus voice, that the need to add an exogenous fluorogenic small molecule might limit application in more complex systems. Stored copy is the green open-access accepted manuscript from UCL Discovery; the published version is Springer Nature text-and-data-mining only, so no figure may be reused.
Lipid droplet dysmetabolism affects cell homeostasis in an in vitro model of Alzheimer’s disease
Applied splitFAST to link lipid-droplet dysmetabolism to cell-homeostasis defects in an in-vitro Alzheimer's model.
Cell-surface receptor-mediated regulation of synaptic organelle distribution controls dendritic spine maturation
Used chemogenetic FAST reporters to study how cell-surface receptors position the spine apparatus for dendritic-spine maturation.
Hi-TARGET: A fast, efficient and versatile CRISPR type I-B genome editing tool for the thermophilic acetogen Thermoanaerobacter kivui
Built a CRISPR type I-B genome-editing toolkit for the thermophilic acetogen Thermoanaerobacter kivui, with pFAST as a fluorescent reporter.
Fluorescence Lifetime Multiplexing with Fluorogen-Activating FAST Protein Variants and Red-Shifted Fluorogens
The 2025 follow-up to the group's 2024 lifetime-multiplexing paper at row 49, moved to a red-shifted fluorogen. FAST mutants that look identical in steady state but differ in lifetime let three structures be separated at once in live cells on one fluorogen, N871b, with H2B and vimentin as the targets.
A tunable and versatile chemogenetic near-infrared fluorescent reporter
Introduced nirFAST, a bright 14-kDa near-infrared reporter for deep, multiplexed imaging that also doubles as a CATCHFIRE proximity tool.
Experimental Approaches to Visualize Effector Protein Translocation During Host-Pathogen Interactions
Names an application nobody has taken up. FAST's potential for studying bacterial effector translocation was shown by pre-labelling a Shigella T3SS effector inside the bacterium before it entered the host cell, and the authors state that studies extending that feature are pending. They also propose pairing FAST with SNAP-tag to follow several effectors at once.
Non-Canonical TERT Activity Initiates Osteogenesis in Calcific Aortic Valve Disease
Cardiovascular disease enters the corpus, and splitFAST is there as a screening instrument rather than as a picture. The authors show that telomerase reverse transcriptase acts outside its canonical role to drive osteogenic reprogramming in human aortic valve interstitial cells, and they read the TERT to STAT5A interaction by fusing the two proteins to NFAST and CFAST and adding HBR-3,5DOM at 5 micromolar. The signal is highly elevated in calcifying cells from day three of osteogenic induction. Their stated reason for choosing the system is the useful part for us: it gives a scalable model that can be used to screen for compounds blocking the TERT to STAT5 interaction. Free full text through PubMed Central, PMC11825275, the publisher version being closed. **Generation, checked 28/10 after the Posts session raised it: this is the original 2019 splitFAST, not splitFAST2.** They cite Tebo and Gautier 2019, name the fragments plainly NFAST and CFAST, and there is no RspA, no splitFAST2 and no Rakotoarison anywhere in the text. That is worth knowing because of what they say they want next: a scalable screen. Self-complementation of the first generation sits at 3.1 micromolar against 220 for splitFAST2, which is the background a compound screen can least afford.
Nano-frFAST: Design of a New Genetically-Encoded Far-Red Fluorescent Label
Designed nano-frFAST, a small far-red fluorogen-activating protein, with a matching series of extended-pi fluorogens.
Synthetically programmed antioxidant delivery by a domesticated skin commensal
Engineered a skin commensal bacterium for programmed antioxidant delivery, using FAST to fluorescently track the strains.
A rapid genome-proteome approach to identify rate-limiting steps in the butyrate production pathway in probiotic <i>Clostridium butyricum</i>, CBM588
A genome-proteome approach to find rate-limiting steps in butyrate production by Clostridium butyricum, with FAST reporting.
Deletion of atypical type II restriction genes in Clostridium cellulovorans using a Cas9-based gene editing system
Improved genetic accessibility of the biofuel-relevant anaerobe Clostridium cellulovorans, using FAST as an anaerobic reporter.
You better keep an eye on your contacts
A Cell Calcium commentary written about two recent splitFAST papers, so a third party devoting a piece to our technology rather than listing it among others. It is the only source we hold that puts a number on the perturbation question: the original splitFAST pair, self-complementation Kd about 3 uM, stabilises ER to lipid-droplet contacts even without fluorogen, while the low-affinity variant at about 220 uM does not and reports faithfully. Full two-page text held, references included.
Engineering, design and selection of fluorescence-activating proteins for advanced imaging and biosensing
A review of how fluorescence-activating proteins like FAST are engineered, designed and selected for advanced imaging.
Fluorescence-activating and absorption-shifting tag
The encyclopedia reference entry defining FAST, the 14-kDa tag engineered from photoactive yellow protein (the Wikipedia article itself).
Mitophagy and release of mitochondrial derived vesicles in cellular models of inducible unfolded protein accumulation in mitochondrial matrix and intermembrane space.
Studied mitophagy and mitochondrial-derived vesicles in inducible cell models, using FAST-family reporters.
The potential of fluorogenicity for single molecule FRET and DyeCycling
The most useful outside assessment we hold, and it is a critique. It tabulates FAST at 14 kDa, about 3.4 nm, fluorogenic ratio 550, Kd 1.3 x 10-7 M, labelling restricted to the C or N terminus, then concludes that the size and that restriction make it impractical and insensitive for nanometre-distance single-molecule FRET. Quantified, sourced, and unflattering, which is what makes it worth holding.
Multipartite Fluorogenic Sensors for Monitoring Tyrosine Phosphatase Activity
First steps toward genetically encoded FAST-based sensors for protein tyrosine phosphatase activity.
Simultaneous detection of membrane contact dynamics and associated Ca2+ signals by reversible chemogenetic reporters
Built reversible splitFAST reporters that simultaneously visualize membrane contact-site dynamics and the Ca2+ signals they carry.
The genetic architecture of protein interaction affinity and specificity
Used splitFAST to quantify how every mutation in the transcription factor JUN reshapes binding across a protein family, mapping interaction affinity and specificity. **It also contains the quantitative cross-validation of splitFAST that the corpus otherwise lacks**: 21 measurements made by a yeast BindingPCA were confirmed by splitFAST in human HCT116 cells, Pearson R = 0.79 (p = 1.72e-5) on absolute binding scores and R = 0.89 (p = 5.3e-5, n = 13) on scores relative to wild type. A 2025 preprint from the same laboratory repeats the point in one line, that yeast BindingPCA correlates strongly with splitFAST in human cells, which makes this an argument a third party is willing to restate.
Genetic Tagging and Imaging of Proteins with iFAST in Candida albicans
A step-by-step protocol for tagging and imaging proteins with iFAST in Candida albicans, 38 iFAST mentions, from the Van Dijck group at KU Leuven. Its premise is ours: the fluorescent proteins available for this pathogen are GFP-derived and poorly suited to it. It pairs with the fungal-tools review at row 145, by the same group, which is the contextual half of the same body of work.
Imaging and proteomics toolkits for studying organelle contact sites
A review of imaging and proteomics toolkits for organelle contact sites, including splitFAST-based approaches.
A versatile bioluminescent probe with tunable color
A tunable-color bioluminescent probe, from FAST-family chemogenetic reporter engineering.
Design of fully synthetic signal peptide library and its use for enhanced secretory production of recombinant proteins in Corynebacterium glutamicum
Built a synthetic signal-peptide library to boost protein secretion in Corynebacterium glutamicum, using FAST-based secretion reporting.
The use of thermostable fluorescent proteins for live imaging in Sulfolobus acidocaldarius
A hyperthermophilic-archaeon imaging study that does not use FAST but tells its readers two things about it: that the oxygen-independent tag has proven useful for live-cell imaging, citing Hernandez and Costa, and that FAST variants with affinities for different ligands are commercially available. A third party pointing its own field at our catalogue.
Lactate-mediated mixotrophic co-cultivation of Clostridium drakei and recombinant Acetobacterium woodii for autotrophic production of volatile fatty acids
Engineered a Clostridium/Acetobacterium co-culture for autotrophic fatty-acid production, with FAST as a fluorescent strain marker.
Teaching old dogs new tricks: genetic engineering methanogens
A 124-reference review of the genetic toolbox for methanogenic archaea, Methanococcus and Methanosarcina foremost, in which FAST appears among the reporter systems. It is the contextual counterpart to our own archaeal rows: the reason FAST is in that toolbox at all is that methanogens are strict anaerobes and the oxygen-dependent fluorescent proteins do not work there.
FAM122A ensures cell cycle interphase progression and checkpoint control by inhibiting B55α/PP2A through helical motifs
Used splitFAST complementation to characterize how FAM122A binds and inhibits the B55-alpha/PP2A phosphatase to control the cell cycle.
Fluorescence lifetime multiplexing with fluorogen activating protein FAST variants
Baranov-group FAST FLIM multiplexing: FAST mutants sharing a fluorogen but differing in fluorescence lifetime, telling several labels apart by lifetime.
A fluorogenic complementation tool kit for interrogating lipid droplet–organelle interaction
A fluorogenic splitFAST toolkit for detecting lipid-droplet-organelle contact sites at nanometer scale in living cells.
Expression and Subcellular Localization of Lanthipeptides in Human Cells
Expressed and localized lanthipeptides in human cells, using FAST-based fluorescent reporting.
A guide to genetically-encoded redox biosensors: state of the art and opportunities
A review of genetically encoded redox biosensors that covers FAST-based fluorogenic designs.
Multiplexed In Vivo Imaging with Fluorescence Lifetime-Modulating Tags
Introduced FAST variants tuned to distinct fluorescence lifetimes, enabling highly multiplexed in-vivo imaging by lifetime rather than color.
LowTempGAL: a highly responsive low temperature-inducible GAL system in Saccharomyces cerevisiae
A low-temperature-inducible gene-expression system in yeast, read out with FAST-based fluorescence.
A multi-colour fluorogenic tag and its application in Candida albicans
Applied a multi-colour fluorogenic FAST tag in the fungal pathogen Candida albicans.
A Combination of Library Screening and Rational Mutagenesis Expands the Available Color Palette of the Smallest Fluorogen-Activating Protein Tag nanoFAST
Expanded the color palette of nanoFAST, the smallest (98-aa) fluorogen-activating protein, by mutagenesis and screening.
Chemogenetic Tools in Focus: Proximity, Conformation, and Sterics
A review of chemogenetic tools (incl. FAST/CATCHFIRE) organized by proximity, conformation and steric mechanisms.
The multiplicity of thioredoxin systems meets the specific lifestyles of Clostridia
Studied thioredoxin redox systems across Clostridia, using FAST-based anaerobic reporting.
Improving Split Reporters of Protein–Protein Interactions through Orthology-Based Protein Engineering
Improved splitFAST's brightness and performance for detecting protein-protein interactions using orthology-based protein engineering.
Integration of Yeast Episomal/Integrative Plasmid Causes Genotypic and Phenotypic Diversity and Improved Sesquiterpene Production in Metabolically Engineered Saccharomyces cerevisiae
Explored how yeast plasmid integration drives phenotypic diversity, using FAST-based auxin-inducible protein degradation.
Refining and illuminating acetogenic Eubacterium strains for reclassification and metabolic engineering
Reclassified and metabolically engineered acetogenic Eubacterium strains, using FAST for fluorescent visualization.
Transforming chemigenetic bimolecular fluorescence complementation systems into chemical dimerizers
A preprint turning chemigenetic bimolecular fluorescence complementation systems into chemical dimerizers.
Engineering and applications of chemogenetic fluorescent tags for advanced biological imaging
The doctoral thesis behind a run of papers we already hold: Lina El Hajji, Sorbonne, supervised by Arnaud Gautier, defended 12 July 2024. nirFAST dominates it with 140 mentions, ahead of pFAST, frFAST, iFAST and CATCHFIRE, and 35 mentions of biosensors. Worth holding because a thesis states the reasoning and the failed attempts that the papers compress out, across mammalian cells, zebrafish and chick.
Inter-organelle contact sites: molecular modulation and generation of new probes to study their dynamics and associated signaling
A study of inter-organelle contact sites introducing new splitFAST-related probes to visualize them.
Effeсts of the 70th Amino Acid Residue on the Photostability of FAST Complexes
A FAST T70V mutant showing the 70th residue barely affects brightness or photostability.
Anaerobic fluorescent reporters for live imaging of Pseudomonas aeruginosa
Developed anaerobic FAST reporters for live imaging of Pseudomonas aeruginosa in biofilms and low-oxygen conditions.
Engineering of Tunable Allosteric-like Fluorogenic Protein Sensors
Engineered tunable allosteric-like FAST biosensors that switch fluorescence in response to a target biomolecule.
Microbiome imaging goes à la carte: Incorporating click chemistry into the fluorescence-activating and absorption-shifting tag (FAST) imaging platform
Added click chemistry to FAST imaging to label anaerobic microbiome bacteria in biofilm communities.
A fluorogenic chemically induced dimerization technology for controlling, imaging and sensing protein proximity
Introduced CATCHFIRE, a small-molecule 'match' that both induces protein proximity and lights it up, for controlling, imaging and sensing protein assembly, transport and interactions.
Fluorogenic chemically induced dimerization
A News & Views highlighting CATCHFIRE, the first chemically induced dimerizer that fluoresces on dimerization.
A fluorescent reporter system for anaerobic thermophiles
Demonstrated a FAST-based reporter that works in anaerobic thermophiles, where oxygen-dependent GFP fails.
Endogenous CRISPR/Cas systems for genome engineering in the acetogens Acetobacterium woodii and Clostridium autoethanogenum
Used endogenous CRISPR/Cas systems to engineer acetogens, with FAST for fluorescent readout.
Application of the Fluorescence-Activating and Absorption-Shifting Tag (FAST) for Flow Cytometry in Methanogenic Archaea
Applied FAST to flow-cytometry analysis and imaging of methanogenic archaea (M. maripaludis, M. acetivorans).
Fluorescence-Activating and Absorption-Shifting Nanoprobes for Anaerobic Tracking
FAST absorption-shifting nanoprobes for tracking anaerobic bacteria.
Production of propionate using metabolically engineered strains of Clostridium saccharoperbutylacetonicum
Engineered Clostridium saccharoperbutylacetonicum for propionate production, tracked with FAST.
Fluorescence-Activating and Absorption-Shifting Tags for Advanced Imaging and Biosensing
An Accounts review by the FAST inventor summarizing how FAST-family tags and biosensors work and their imaging and biosensing applications.
Multifunctional stimuli-responsive chemogenetic platform for conditional multicolor cell-selective labeling
Engineering on the fluorogen side rather than the protein side: stimuli-responsive fluorogenic ligands are paired with organelle-targeted FAST so that labelling becomes conditional on a stimulus, giving multicolour cell-selective labelling. 76 FAST mentions and the authors name the platform srFAST. A third group building a product on top of our tag, which is the kind of use worth knowing about.
Spatial Structure of NanoFAST in the Apo State and in Complex with its Fluorogen HBR-DOM2
A solution-NMR answer to why the smallest FAST variant is also the fussiest. Truncating FAST leaves the remaining fold intact, but the deleted N-terminus had been stabilising the C-terminal domain in the apo state, which is why nanoFAST works well with only one of the known fluorogens, HBR-DOM2. Structural grounding for the ligand-promiscuity limit, not a new tag.
Cyanine Dyes as Fluorogens for FAST and NanoFAST Proteins
A fluorogen-chemistry survey outside the usual rhodanine family: seven new cyanine dyes synthesised and eighteen in all probed against FAST and nanoFAST, to find which structural elements make a workable cyanine-based pair. Useful as the record of what has already been tried and did not become a product.
The Fluorescence-Activating and Absorption-Shifting Tag (FAST) Enables Live-Cell Fluorescence Imaging of Methanococcus maripaludis
Showed FAST enables live-cell fluorescence imaging of the methanogenic archaeon Methanococcus maripaludis under anoxic conditions.
Structure-based rational design of an enhanced fluorogen-activating protein for fluorogens based on GFP chromophore
Rational engineering of FAST rather than screening. From the solution structure of the FAST/N871b complex, NMR relaxation and simulation, the authors locate the mobile regions that cost quantum yield and design mutants optimised for that one fluorogen. N871b is the same red-shifted fluorogen the group's FLIM multiplexing work at row 143 later builds on, so the two sit together.
Encoding with a fluorescence-activating and absorption-shifting tag generates living bacterial probes for mammalian microbiota imaging
Encoded gut-microbiota bacteria with FAST to create living probes for imaging the mammalian microbiota.
Isolating and Engineering Fluorescence-Activating Proteins Using Yeast Surface Display
A protocol for isolating and engineering new fluorescence-activating proteins by yeast surface display.
Autotrophic lactate production from H2 + CO2 using recombinant and fluorescent FAST-tagged Acetobacterium woodii strains
Engineered fluorescent FAST-tagged Acetobacterium woodii for autotrophic lactate production from H2 + CO2.
Establishment of Green- and Red-Fluorescent Reporter Proteins Based on the Fluorescence-Activating and Absorption-Shifting Tag for Use in Acetogenic and Solventogenic Anaerobes
Established greenFAST and redFAST reporters for use in acetogenic and solventogenic anaerobes.
Recent Advancements in Tracking Bacterial Effector Protein Translocation
A review of methods for tracking bacterial effector-protein translocation, featuring FAST among the reporters.
Practical observations on the use of fluorescent reporter systems in Clostridioides difficile
Practical comparison of fluorescent reporter systems in Clostridioides difficile: CFPopt, mCherryOpt, phiLOV2.1, SNAPCd and HaloTag, concluding that SNAPCd performs best. FAST is NOT used here: it appears only in the discussion, as an untested option still requiring a substrate. Mention only.
New Tools for Anaerobic Bacteria: Metabolically Engineered Strains to Improve Energy Supply, Recombinant Production of Value-Added Products, and Fluorescent Reporter Proteins - ProQuest
A web resource on metabolically engineered anaerobic strains (low-value, non-peer-reviewed item).
Engineering of a fluorescent chemogenetic reporter with tunable color for advanced live-cell imaging
Introduced pFAST, a promiscuous FAST that binds a whole palette of fluorogens, letting one tag be imaged in tunable colors across the visible spectrum.
Transient Fluorescence Labeling: Low Affinity—High Benefits
A review whose thesis is the FAST argument stated as a principle: low binding affinity is a feature, because transient labelling is what makes single-molecule localisation and exchangeable probes possible. FAST appears eighteen times and HMBR four, alongside nanoFAST, and the malachite-green FAP lineage is treated separately.
Inheritance of the reduced mitochondria of Giardia intestinalis is coupled to the flagellar maturation cycle
Studied inheritance of Giardia's reduced mitochondria (mitosomes), using FAST-based imaging.
Anaerobic fluorescent reporters for cell identification, microbial cell biology and high-throughput screening of microbiota and genomic libraries
A review of real-time fluorescent reporters (FAST prominent among them) for anaerobes in gene expression, screening and cell biology.
Four species of bacteria deterministically assemble to form a stable biofilm in a millifluidic channel
Used FAST to build and image a stable four-species model biofilm in a millifluidic channel.
Protein-fragment complementation assays for large-scale analysis of protein-protein interactions
A review of protein-fragment complementation assays for large-scale PPI analysis, including splitFAST.
ER-mitochondria contact sites in neurodegeneration: genetic screening approaches to investigate novel disease mechanisms
A third party recommending splitFAST rather than describing it. Reviewing the ways to see ER-mitochondria contacts in neurodegeneration, the authors set out why irreversible split reporters fall short and write that one solution would be an adapted system such as splitFAST, whose fragments dissociate when the two membranes separate.
Developing Bright Green Fluorescent Protein (GFP)-like Fluorogens for Live-Cell Imaging with Nonpolar Protein−Chromophore Interactions
Developed brighter GFP-like fluorogens for FAST using nonpolar protein-chromophore interactions.
Production of the biocommodities butanol and acetone from methanol with fluorescent FAST-tagged proteins using metabolically engineered strains of Eubacterium limosum
Produced butanol and acetone from methanol in engineered Eubacterium limosum, tracked with fluorescent FAST tags.
NanoFAST: structure-based design of a small fluorogen-activating protein with only 98 amino acids
Introduced NanoFAST, a structure-based minimal fluorogen-activating protein of just 98 amino acids.
Styrene Derivatives of Indole and Pyranone as Fluorogenic Substrates for FAST Protein
Tested styrene derivatives of indole and pyranone as new fluorogens for FAST.
Fluorescent toys 'n' tools lighting the way in fungal research
A 301-reference survey of the fluorescent toolbox for fungal research, Candida above all with 194 mentions, in which FAST, splitFAST and the HBR fluorogens each appear. From Van Genechten and Van Dijck at KU Leuven, the same group behind the iFAST protocol in Candida albicans, so it is the contextual half of a body of work we already touch through our Candida rows.
Auxin-mediated protein depletion for metabolic engineering in terpene-producing yeast
An auxin-inducible protein-depletion tool for metabolic engineering in terpene-producing yeast, read out with FAST.
Color Tuning of Fluorogens for FAST Fluorogen-Activating Protein
Created a panel of color-shifted benzylidene-imidazolone fluorogens for FAST without losing binding efficiency.
Emergent Biosensing Technologies Based on Fluorescence Spectroscopy and Surface Plasmon Resonance
Four accurate sentences placing our sensing variants among emergent biosensor technologies: cpFAST forming green-yellow complexes with HMBR or orange-red with HBR-3,5-DOM, wired to M13 at one terminus and calmodulin at the other, and split-FAST as a reversible reader of fast Ca2+ transients.
Orthogonal fluorescent chemogenetic reporters for multicolor imaging
Introduced greenFAST and redFAST, spectrally orthogonal FAST variants, for simultaneous two-color imaging in live cells and zebrafish.
Near-infrared fluorescent probes: a next-generation tool for protein-labeling applications
A near-infrared probe review from the Kikuchi lab in Osaka, which places frFAST, named nine times, inside the NIR protein-labelling landscape. Useful mainly as context for where our far-red variant sits: the review's centre of gravity is elsewhere, with sixteen HaloTag and seventeen SNAP-tag mentions, so it is a map of the neighbourhood rather than an assessment of us.
Interspecies Microbial Fusion and Large-Scale Exchange of Cytoplasmic Proteins and RNA in a Syntrophic Clostridium Coculture
Revealed interspecies cytoplasmic exchange in a Clostridium coculture, using FAST-based fluorescent tracking.
Fluorescent secreted bacterial effectors reveal active intravacuolar proliferation of Listeria monocytogenes in epithelial cells
Showed FAST can tag secreted bacterial effectors, revealing Listeria actively proliferating inside epithelial-cell vacuoles in real time.
Development of Strong Anaerobic Fluorescent Reporters for Clostridium acetobutylicum and Clostridium ljungdahlii Using HaloTag and SNAP-tag Proteins
Developed strong anaerobic FAST reporters for Clostridium acetobutylicum and C. ljungdahlii.
Visualizing the dynamics of exported bacterial proteins with the chemogenetic fluorescent reporter FAST
Used FAST to visualize the dynamics of proteins exported to the bacterial cell surface.
A Far-Red Emitting Fluorescent Chemogenetic Reporter for In Vivo Molecular Imaging
Introduced frFAST, a far-red FAST variant, for spectral multiplexing and deep-tissue in-vivo imaging in zebrafish and mammalian cells.
Illuminating Cellular Biochemistry: Fluorogenic Chemogenetic Biosensors for Biological Imaging
A review of fluorogenic chemogenetic biosensors, including FAST-based designs, for reading out cellular biochemistry by fluorescence.
Sensing cellular biochemistry with fluorescent chemical–genetic hybrids
A review arguing that hybrid chemical-genetic reporters like FAST overcome the photostability and brightness limits of fluorescent-protein biosensors.
Next-Generation Fluorogen-Based Reporters and Biosensors for Advanced Bioimaging
A review of next-generation fluorogen-based reporters and biosensors, covering FAST and related chemogenetic systems for advanced bioimaging.
Red-Shifted Substrates for FAST Fluorogen-Activating Protein Based on the GFP-Like Chromophores
Reported a red-fluorescent GFP-like fluorogen for FAST for live-cell microscopy.
A Strongly Fluorescing Anaerobic Reporter and Protein-Tagging System for Clostridium Organisms Based on the Fluorescence-Activating and Absorption-Shifting Tag Protein (FAST)
First strongly fluorescent, oxygen-independent FAST reporter and protein-tagging system for anaerobic Clostridium.
A split fluorescent reporter with rapid and reversible complementation
Introduced splitFAST, FAST split into two fragments that fluoresce only when reunited, giving a rapid, reversible readout of transient protein-protein interactions in live cells.
Single-Molecule Localization Microscopy with the Fluorescence-Activating and Absorption-Shifting Tag (FAST) System
Demonstrated FAST for single-molecule localization super-resolution microscopy, exploiting its fast fluorogen on/off binding to pinpoint molecules.
Live cell super resolution imaging by radial fluctuations using fluorogen binding tags
Applied FAST to super-resolution radial-fluctuation (SRRF) imaging, exploiting its stochastic fluorogen binding.
Improved Chemical-Genetic Fluorescent Markers for Live Cell Microscopy
Introduced improved FAST variants (iFAST and tandem tdFAST) as brighter chemical-genetic markers for live-cell microscopy.
Fluorogenic Protein-Based Strategies for Detection, Actuation, and Sensing
A BioEssays review by the FAST inventors on fluorogenic protein-based strategies for detection, actuation and sensing; a strong secondary source for Wikipedia.
Circularly Permuted Fluorogenic Proteins for the Design of Modular Biosensors
The paper that introduced cpFAST, and it had been missing from the KB while six rows carried the reporter it created. Circular permutation makes fluorogen binding conditional on analyte recognition, which turns FAST from a label into a sensing module; they prove it with plug-and-play multicolour Ca2+ biosensors in live mammalian cells, calmodulin and M13 providing the recognition.
Engineering <i>Clostridium</i> organisms as microbial cell-factories: challenges & opportunities
A review of engineering Clostridium as microbial cell factories, where FAST features as an anaerobic reporter.
The inducible chemical-genetic fluorescent marker FAST outperforms classical fluorescent proteins in the quantitative reporting of bacterial biofilm dynamics
Showed the inducible FAST marker outperforms classical GFP-family reporters for quantifying bacterial biofilm dynamics.
Fluorogenic Probing of Membrane Protein Trafficking
Used FAST with a non-permeant fluorogen to selectively image cell-surface versus intracellular proteins, tracking membrane-protein trafficking.
High-throughput, image-based screening of pooled genetic-variant libraries
A high-throughput image-based screening method for pooled genetic-variant libraries, with FAST-relevant readout.
Chromophore Renewal and Fluorogen-Binding Tags: A Match Made to Last
The paper behind the photobleaching-resistance argument, and it is more careful than the slogan. Reversible fluorogen binding does cut the apparent photobleaching rate of Y-FAST sharply, because a bleached fluorogen is simply exchanged for a fresh one. But the authors then show where the limit sits: photostability is capped not by the dye but by oxidation of specific residues in the protein scaffold, by reactive oxygen species the bound fluorogen itself generates. They also give a generic experimental and theoretical method for measuring that, which had not existed.
Fluorogenic Labeling Strategies for Biological Imaging
A review of fluorogenic labeling strategies for lighting up proteins and RNA in live cells, situating FAST among the emerging chemogenetic tools.
Dynamic multicolor protein labeling in living cells
Showed FAST can be imaged in several colors by swapping fluorogens, enabling dynamic multicolor labeling of proteins in living cells.
Small-Molecule-Induced and Cooperative Enzyme Assembly on a 14-3-3 Scaffold
Studied small-molecule-induced enzyme assembly on a 14-3-3 scaffold; early induced-proximity work relevant to CATCHFIRE.
Development of bioorthogonal fluorogenic reporters for biological imaging
Chenge Li's doctoral thesis, ENS and PSL, supervised by Arnaud Gautier, with Ludovic Jullien on the jury. The densest single document we hold on the early fluorogen chemistry: 235 mentions of FAST, 162 of HBR and 69 of HMBR. This is where the rhodanine series that became the product line was worked out, including what was tried and set aside.
Shedding light on biology of bacterial cells
Our advantages enumerated by an outside review in 2016, before most of our own literature existed: about 14 kDa and monomeric, a non-toxic cell-permeable fluorogen, binding that is instantaneous on the millisecond scale, highly specific and fully reversible, a quantum-yield gain with an absorption red shift, and a forward look to blinking behaviour for super-resolution.
Small fluorescence-activating and absorption-shifting tag for tunable protein imaging in vivo
The founding FAST paper: a 14-kDa tag, half the size of GFP, that fluoresces only while reversibly bound to its fluorogen, giving wash-free, tunable, reversible protein labeling.
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