Ratiometric Fluorescence Changes Research Articles

Ratiometric Normalization of Near-Infrared Fluorescence in Defect-Engineered Single-Walled Carbon Nanotubes for Cholesterol Detection.

Ratiometric probing of analytes presents a substantial advancement in molecular recognition, offering self-calibrating signals that enhance the measurement accuracy and reliability. We present a dual-emitting probe based on (6,5) chirality-enriched single-walled carbon nanotubes (SWCNTs) with oxygen defects for cholesterol (Chol) detection using ratiometric fluorescence readouts. The interaction with Chol induced significant intensity variations in the E11 and E11* emission peaks of oxygen defect-induced SWCNTs, giving rise to ratiometric fluorescence changes. The sensitivity of these probes toward Chol in water and serum was 0.28 ± 0.01 and 0.72 ± 0.05 μM, respectively, which is comparable to that of common gold standards for cholesterol detection used in clinical samples. By utilizing ratiometric readouts, our approach enhanced selectivity over numerous competing analytes, including amino acids, sugars, cations, anions, proteins, steroid hormones, surfactants, and phospholipids. Mechanistic investigations revealed that Chol detection by defect-integrated SWCNTs was facilitated by Chol incorporation within micelles formed by sodium cholate, the surfactant dispersant used for the SWCNT suspension. Oxygen defects played a crucial role by directly interacting with Chol. This strategy employing defect-integrated dual-peak NIR-emitting SWCNTs as sensors for Chol in aqueous and serum environments not only enables background-free detection of biologically relevant analytes but also advances biosensing using SWCNTs through tailored surface functionalization and advanced read-out concepts.

Novel Colorimetric and Near-Infrared Ratiometric Fluorescent Probe for Sensing Cysteine in Food Samples, Plants, and Living Cells.

Cysteine (Cys) is a crucial biothiol that acts a significant function in food samples and biological systems, including plant roots and living cells. Hence, we developed a novel colorimetric and near-infrared ratiometric fluorescent probe (CT), composed of coumarin and tetrahydroacridine-conjugated indole salt, for the detection of Cys. Upon reaction with Cys, the probe undergoes a specific N-substitution reaction, resulting in a notable colorimetric change and a significant ratiometric fluorescent response in both visible and near-infrared emission channels. These dual-channel ratiometric fluorescence changes are completely independent, enabling the probe to obtain great selectivity, sensitivity, and exceptional detection accuracy. Leveraging these attributes, the probe was employed to provide accurate quantitative analysis of Cys in food samples. Furthermore, confocal imaging demonstrated that the probe could monitor both exogenous and endogenous Cys levels in living cells and track Cys changes in plant roots under heavy metal stress. This work presents a dependable and accurate imaging solution for tracking and identifying Cys of real food, plants, and living cells.

High-performance ratiometric fluorescence detection and removal of tetracycline in milk based on CDs@ZSM-5:Eu3+

Exploring materials with the dual functionality of detecting and removing tetracycline (TC) residues is crucial because of the environmental and health risks posed by antibiotic overuse. This study introduces a dual-emissive luminescent probe, CDs@ZSM-5:Eu3+, created through a solvent-free method combined with subsequent Eu3+ion exchange. The nanocomposite's blue emission, originating from carbon dots (CDs), is quenched by TC via an internal filtering effect, while an antenna effect triggers a strong red fluorescence of a TC-Eu3+chelate. The ratiometric fluorescence changes in CDs@ZSM-5:Eu3+ endow a self-calibrated sensing mechanism for TC, offering a low detection limit of 5.04 nM and a broad detection range of 0.01–50 μM. Demonstrated in real milk samples, the probe exhibits high selectivity and accuracy in detecting TC. The nanocomposite also displayed an impressive TC removal capacity of 238.1 mg g−1 in water, ascribing to the enrichment and electrostatic attraction effects of ZSM-5 toward TC molecules. This research offers a facile strategy for constructing multifunctional zeolite-based hybrids for simultaneous TC detection and removal from aqueous solutions.

Self-correcting ratiometric fluorescence assay for H2S detection in live cells and releasing agents

Hydrogen sulfide (H2S) is both a biologically significant signaling molecule, and a harmful gas that is generated in the environment; thus, its detection and monitoring are required in range of biological and environmental systems. Most fluorescent probes for H2S detection rely on single-detection fluorescence signals, making them vulnerable to interference from other species in complex microenvironment. In this study, we developed a ratiometric fluorogenic and colorimetric assay for H2S detection, in which pyrene and naphthalimide fluorophores operate together to provide a selective and sensitive detection system. Pyrene maintains fluorescence (always ‘On’), while naphthalimide exhibits an ‘On-Off’ response through thiolysis with H2S. By measuring the ratiometric fluorescence from these moieties, a self-calibrating response can be measured that is resistant to interference from other species in the microenvironments. The probe was used to measure H2S generation from H2S donor substrates and to monitor H2S fluctuations within living cells through selective, ratiometric fluorescence changes and real-time, dual-channel fluorescence imaging.

Dehydro acetic acid analogue Schiff base as multimode fluorescence chemosensors for Th4+ and Ni2+ ions and its application to in-vivo bioimaging

Dehydroacetic acid analogue (DHA) and 7-(diethylamino)-2-oxo-2H-chromene-3-carbohydrazide (COM) Schiff base ligand (COM-DHA)as dual-mode fluorescence chemosensorwas synthesized and structurally characterised by using elemental analysis, FTIR, NMR, and Mass analysis. The free probe COM-DHA shows distinct fluorescence emission at 476 nm, upon interaction with Ni2+ the emission intensity was quenched and with Th4+ the emission intensity was redshirted to 500 nm. Further, the ratiometric fluorescence change (I500/I476) was used to detect Th4+, and a turn-off fluorescence probe COM-DHA at 476 nm was used to detect Ni2+ ions. The COM-DHA forms a 2:1 stoichiometric complex with both Ni2+/Th4+ ions with an estimated association constant of 4.84 × 104 and 6.68 × 104 M−2 respectively. A further probe COM-DHA is that it has a wide working pH range (5 to 10) with a short response time (2 to 3 min). The detection limit of COM-DHA towards Ni2+ and Th4+ was found to be 77.8and 48.7 nM, respectively. The, DFT/TD-DFT calculations and FTIR analysis were used to study the binding mechanism between COM-DAH and Ni2+/Th4+ ions. Furthermore, the proposed probe COM-DHAhas successfully quantified trace amounts of Ni2+ and Th4+in real samples and applied them to the in-vivo bioimaging of Th4+ in C. elegans model.

Visible light-triggered NIR ratiometric fluorescent metal-free CO-releasing molecule for self-monitoring of CO delivery and effective cancer therapy

Cancer is a serious global health issue, and exploring effective treatment methods is of great significance for cancer prevention and control. Carbon monoxide (CO), as an important gas signaling molecule in the life system, has been proven to have good anti-cancer effects. However, how to controllably, safely, and effectively deliver CO to the tumor site for clinical treatment remains a challenge. Herein, a new metal-free CO-releasing molecule COR-XAC was developed for controlling CO release and cancer therapy. COR-XAC is based on the hybrid of 3-hydroxyl flavone and oxanthracene fluorophores, showing visible light-controlled CO-releasing properties and near-infrared (NIR) ratiometric fluorescence changes at 690 and 760 nm. COR-XAC shows low cytotoxicity and can be successfully applied to release CO in cells and tumors, and the CO-releasing and delivery process could be monitored by its own NIR ratiometric fluorescence changes. More importantly, the anti-cancer performance of COR-XAC was evaluated in 4T1 tumor mice, and it was found that COR-XAC plus light illumination showed excellent tumor inhibition effect, which provided a promising new effective method for cancer treatment.

A Ratiometric Benzimidazole-Based Fluorescent Probe for The Recognition of Phosgene in Solution and Gaseous Phases.

Considering the high toxicity and widespread application of phosgene, there is an urgent need to develop a simple and sensitive method for detecting phosgene. In this work, we designed and synthesized a novel ratiometric fluorescent probe 1 containing fluorophores of benzimidazole and benzothiazole. Probe 1 showed excellent sensitivity (< 30s) and selectivity (LOD = 3.82 nM) for phosgene and significant ratiometric fluorescence changes. In addition, 1-loaded polystyrene membrane test strips were used to conveniently and efficiently detect phosgene gas (0.5 ppm) via the naked eye and the RGB APP of the smartphone, indicating that this probe has great potential for phosgene detection in the gaseous phase.

Red-emitting ratiometric fluorescent metal-free CO-releasing molecule triggered by visible light for controllable and trackable CO delivery and efficient treatment of cutaneous wounds

CO is a highly toxic gas but also an important gasotransmitter in the human body that regulates cell functions and shows great therapeutic effects in clinical trials. Due to the unsafe nature of directly inhaling CO gas, the controllable and trackable release and delivery of CO are currently the key and main challenges in achieving CO therapy. In this work, we developed a novel red-emitting fluorescent metal-free CO-releasing molecule (CORM) named COR-FC that can be used to release CO under visible light irradiation and deliver CO in cells safely, accompanied by ratiometric fluorescence changes. COR-FC combines coumarin and 3-hydroxy flavonoid fluorophores and shows low cytotoxicity. Under the light irradiation of 460 nm, COR-FC undergoes rapid photolysis to release CO, accompanied by a decrease in fluorescence at 630 nm and a significant enhancement in fluorescence at 520 nm. More importantly, COR-FC can release CO in cells and tissues under visible light illumination, which provides a new metal-free CORM achieving precise control with self-fluorescence tracking. The released CO showed good anti-inflammatory effects and was successfully used to promote wound healing, indicating that COR-FC has the potential to be further applied in the biomedical field.

An AIE and ESIPT based ratiometric fluorescent hydrogel sensor for detecting biogenic amines and chicken breast freshness

Meat quality is closely linked to food safety and human health, with freshness being a key indicator. Therefore, it is crucial to explore reliable and effective methods for detecting meat freshness. This study presented a ratiometric fluorescent hydrogel sensor based on aggregation-induced emission (AIE) and excited state intramolecular proton transfer (ESIPT) for this purpose. Tetraphenylene (TPE) and 2-(2′-hydroxyphenyl)-benzothiazole (HBT) fluorophore were intersected to generate TPE-HBT, and then a novel biogenic amines (BAs) detection ratiometric fluorescent probe, TPE-HBTAc, was successfully prepared by hydroxyl acetylation. TPE-HBTAc displayed a large Stokes shift and good photostability. It showed a significant response to the representative biogenic amine trimethylamine (TMA), with a ratiometric fluorescence change and low detection limit (LOD = 65 nM). Furthermore, BAs detection hydrogel sensor was created by mixing the fluorescent probe TPE-HBTAc in an agarose hydrogel with a 3D polymer network. The hydrogel sensor displayed a distinct color change from blue to yellow when exposed to TMA vapor under UV light and had a low detection limit (LOD = 5.88 ppm). The hydrogel sensor was utilized in conjunction with a smartphone RGB analysis system to determine the freshness of chicken breast samples. Consumers can easily observe the chicken breast freshness in real-time and non-destructively through UV flashlight illumination. The results of this work suggest that ratiometric fluorescent hydrogel sensor based on AIE and ESIPT can serve as a promising strategy to monitor food freshness.

Swift and quantitative naked-eye detection of cyanide: A dual-mode approach using an ICT-based colorimetric and ratiometric fluorescent chemodosimeter

A chemodosimeter (1) based on dicyanovinyl with a strong intramolecular charge transfer (ICT) mechanism has been devised for the selective and sensitive detection of cyanide (CN−) in a tetrahydrofuran aqueous solution. In polar solvents, chemodosimeter 1 exhibits robust ICT-induced emission; however, in the solid state, its fluorescence weakens owing to the formation of molecular aggregates prompted by intermolecular π–π interactions. The colorimetric and ratiometric fluorescent changes observed in the chemodosimeter upon exposure to CN− result from the incorporation of CN− at the β-conjugated site of the dicyanovinyl moiety of chemodosimeter 1, effectively hindering the ICT process. This alteration is supported by 1H NMR titration experiments and density functional theory calculations, which corroborate the cyanide-induced ICT on–off switching mechanism. The detection limit of chemodosimeter 1 (5.8 × 10−7 M) for CN− falls below the maximum allowable level of cyanide in drinking water (1.9 × 10−6 M), as established by the World Health Organization. Test strips utilizing chemodosimeter 1 offer a potential solution for swift and quantitative naked-eye detection of CN− in real-time scenarios. Moreover, chemodosimeter 1 was effectively employed for detecting cyanide in actual environmental water samples.

Construction of an ultrasensitive hypochlorite fluorescent probe based on a novel chromophore fragmentation strategy

The excess intake of hypochlorite (ClO−) in the human body and excessive emission in wastewater can pose serious health risks and environmental pollution. Therefore, it is vital to develop an efficient method for detecting ClO− in foodstuff and water. In this work, we reported a lactam-fused aza-BODIPY (LAB1) for ultrasensitive ClO− detection based on novel chromophore fragmentation strategy for the first time. It exhibited distinct advantages of good selectivity, high sensitivity, fast response (60 s) and low detection limit of 9.65 nM. The sensing mechanism revealed LAB1 core was fragmented by ClO−, leading to significantly colorimetric and ratiometric fluorescence changes. As a result, 320 nm blue-shift in absorption maximum (from green to colorless) and 340 nm blue-shift in emission maximum (from dark to bright blue) allowed for visual detection by the naked eyes. The potential applicability of LAB1 for ClO− detection with satisfactory recovery in the real samples (tap water and lake water) and vegetables (spinach, lettuce, Chinese cabbage, bean sprouts), and fluorescence “turn on” imaging in mung bean sprout were demonstrated. In addition, LAB1-loaded test paper was suitable for semi-quantitative scanning ClO− analysis by using color recognition APP in a smartphone. Therefore, this work highlights a promising approach based on novel sensing mechanism for monitoring ClO− in food samples.

Ratiometric fluoroprobe based on Eu-MOF@Tb3+ for detecting tetracycline hydrochloride in freshwater fish and its application in rapid visual detection

Tetracycline hydrochloride (TCH), a prevalent antibiotic in aquaculture for treating bacterial infections, poses challenges for on-site detection. This study employed the reversed-phase microemulsion method to synthesize a uniform nano metal-organic framework (MOF) material, europium-benzene-p-dicarboxylic acid (Eu-BDC), doped with Tb3+ to form a dual-emission fluorescence probe. By leveraging the combined a-photoinduced electron-transfer (a-PET) and inner filter effect (IFE) mechanisms, high-sensitivity TCH detection in Carassius auratus and Ruditapes philippinarum was achieved. The detection range for TCH is 0.380–75 μM, with a low limit of detection (LOD) at 0.115 μM. Upon TCH binding, Eu-BDC fluorescence rapidly decreased, while Tb3+ fluorescence remained constant, establishing a ratiometric fluorescence change. Investigation into the TCH quenching mechanism on Eu-BDC was conducted using time-dependent density functional theory (TD-DFT) calculations and fluorescence quenching kinetic equations, suggesting a mixed quenching mechanism. Furthermore, a novel photoelectric conversion fluorescence detection device (FL-2) was developed and evaluated in conjunction with high-performance liquid chromatography-diode-array detection (HPLC-DAD). This is the first dedicated fluorescence device for TCH detection, showcasing superior photoelectric conversion performance and stability that reduces experimental errors associated with smartphone photography methods, presenting a promising avenue for on-site rapid TCH detection.

Aptamer Conjugate-Based Ratiometric Fluorescent Probe for Precise Imaging of Therapy-Induced Cancer Senescence.

Therapy-induced cellular senescence has been increasingly recognized as a key mechanism to promote various aspects of carcinogenesis in a nonautonomous manner. Thus, real-time imaging monitoring of cellular senescence during cancer therapy is imperative not only to further elucidate its roles in cancer progression but also to provide guidance for medical management of cancer. However, it has long been a challenging task due to the lack of effective imaging molecule tools with high specificity and accuracy toward cancer senescence. Herein, we report the rational design, synthesis, and evaluation of an aptamer conjugate-based ratiometric fluorescent probe for precise imaging of therapy-induced cancer senescence. Unlike traditional senescence imaging systems, our probe targets two senescence-associated markers at both cellular and subcellular dimensions, namely, aptamer-mediated membrane marker recognition for active cell targeting and lysosomal marker-triggered ratiometric fluorescence changes of two cyanine dyes for site-specific, high-contrast imaging. Moreover, such a two-channel fluorescence response is activated after a one-step reaction and at the same location, avoiding the diffusion-caused signal decay previously encountered in dual-marker activated probes, contributing to spatiotemporally specific imaging of therapy-induced cancer senescence in living cells and three-dimensional multicellular tumor spheroids. This work may offer a valuable tool for a basic understanding of cellular senescence in cancer biology and interventions.

Construction of a ratiometric fluorescent probe for discriminative detection of SO2 derivatives and Cys and its bio-imaging application

Endogenous sulfur dioxide (SO2) and cysteine (Cys) are important species and interacted with each other. However, due to the reactivity of these species, it is difficult to differentiated detection them. Thus, it is necessary to develop new fluorescent probe for discriminative detect these reactive sulfur species in biological environment. Herein, A new hemicyanine derivative Mito-AE with dual recognition sites for SO2 derivatives and Cys detection was constructed and applied in biological detection. Mito-AE exhibited ratiometric fluorescence change (F470/F610) of over 90-fold when interacted with SO2, and a dramatic fluorescence turn-on at 610 nm when interacted with Cys. The probe can monitor both SO2 derivatives and Cys with fast response rate (within 1 min), high sensitivity and selectivity. As proved by biological experiments, Mito-AE was able to successfully detect exogenous and endogenous SO2 in living cells as well as in mitochondria area. More importantly, the detection of SO2 derivatives and Cys in zebrafish model suggesting a potential application for studying the multiple interaction of biological sulfurous species in live organism.

An AIE-based ratiometric fluorescent probe for reversible detection of bisulfite and hydrogen peroxide and its application in food samples and biological imaging

Reversible detection of bisulfite (HSO3-) and hydrogen peroxide (H2O2) is highly desirable in biology and food safety owing to their redox pair function in biological systems and their utilization in food industry. In this work, a ratiometric fluorescent probe (TPA-Y) with aggregation-induced emission (AIE) properties based on triphenylamine fluorophore was designed and synthesized for reversibly monitoring of HSO3-/H2O2. Reaction of HSO3- with TPA-Y can result in ratiometric fluorescence changes (I444/I629), and the emission spectrum blue shift reaches 185 nm, which can effectively avoid spectral overlap. Meanwhile, the solution color changed obviously from orange to colorless, enabling naked eye detection of HSO3-. TPA-Y was capable of quantitatively detect HSO3- in sugar and wine samples with the recovery rates ranging from 94.52 to 102.18%. In addition, TPA-Y is suitable to prepare paper strips and anti-counterfeiting ink for HSO3- detection. In addition, the original TPA-Y can be released after the addition product (TPA-Y-HSO3-) is oxidized by H2O2, and the HSO3-/H2O2 redox cycle can repeat for at least six times. The probe TPA-Y can be applied to fluorescence imaging of reversible and dynamic interconversion between HSO3- and H2O2 in living MCF-7 cells.

A ratiometric fluorescent probe based on cyclization mechanism to identify hydrazine in environment and living cells

A colorimetric and ratiometric fluorescent probe containing hydroxyl-chalcone moiety as sensing site has been investigated to monitor hydrazine in actual water/soils, gas environment, and living cells. After adding hydrazine, the chalcone structure was cyclized and dihydropyrazole ring was formed, realizing the obvious fluorescence change from red (590 nm) to blue (480 nm) with 110 nm fluorescence peak shift. The probe has low detection limit (0.45 µM), large upgraded ratiometric fluorescence change (I480/I590, 66-fold), and wide pH work range (3.0-10.0). Probe CCA also can monitor hydrazine from actual water/soil samples, liquid/vapor hydrazine with portability and living cells.

A terpyridyl-rhodamine hybrid fluorescent probe for discriminative sensing of Hg (II) and Cu (II) in water and applications for molecular logic gate and cell imaging

Sensitive and discriminative sensing of more than one analyte with a single fluorescent probe is significant and challenging. Herein a new terpyridyl-rhodamine hybrid, namely TRH, has been rationally designed and prepared with two responsive groups in the molecular structure, which facilitate the discriminative detection of Hg2+ and Cu2+ ions in water with detection limits of 4.9 and 53.3 nM by ratiometric fluorescence change (F595/F485) and fluorescence quenching, respectively. Investigations show that the selectivity to Hg2+ ions can be attributed to Hg2+-promoted spirolactam ring opening and further hydrolysis, followed by a through-bond energy transfer (TBET) process. The selective fluorescence quenching to Cu2+ ions probably can be ascribed to the binding Cu2+ to terpyridyl that triggers a ligand-to-metal charge transfer (LMCT) process, which can also efficiently inhibit the TBET process induced by Hg2+ ions and promotes the discriminative sensing of Cu (II) and Hg (II). In addition, the fluorescent responses to Hg2+ and Cu2+ ions cover a wide pH range. Moreover, a combinatorial logic gate with the functions of NOR and INHIBIT has been fabricated by using Hg2+ and Cu2+ ions as chemical input signals, and fluorescence at 485 and 595 nm as output signals. Besides, TRH also exhibits sensitive and discriminative sensing ability to Hg2+ and Cu2+ ions by the fluorescence of rhodamine fluorophore. Significantly, based on the fluorescence signal output of rhodamine moiety, TRH can be used as a tracer for the discriminative sensing of Hg2+ and Cu2+ ions by using living cells.

Ultrathin C3N4 nanosheets-based oxidase-like 2D fluorescence nanozyme for dual-mode detection of organophosphorus pesticides

Engineering efficient dual-mode portable sensor with built-in cross reference correction is of great significance for onsite reliable and precise detection of organophosphorus pesticides (OPs) and evading the false-positive outputs, especially in emergency case. Currently, most nanozyme-based sensors for OPs monitoring primarily replied on the peroxidase-like activity, which involved unstable and toxic H2O2. In this scenario, a hybrid oxidase-like 2D fluorescence nanozyme (PtPdNPs@g-C3N4) was yielded by in situ growing PtPdNPs in the ultrathin two-dimensional (2D) graphitic carbon nitride (g-C3N4) nanosheet. When acetylcholinesterase (AChE) hydrolyzed acetylthiocholine (ATCh) to thiocholine (TCh), it ablated O2−• from the dissolved O2 catalyzed by PtPdNPs@g-C3N4’s oxidase-like activity, hampering the oxidation of o-phenylenediamine (OPD) into 2,3-diaminophenothiazine (DAP). Consequently, with the increasing concentration of OPs which inhibited the blocking effect by inactivating AChE, the produced DAP caused an apparent color change and a dual-color ratiometric fluorescence change in the response system. Through integrating into a smartphone, a H2O2-free 2D nanozyme-based onsite colorimetric and fluorescence dual-mode visual imaging sensor for OPs was proposed with acceptable results in real samples, which holds vast promise for further development of commercial point-of-care testing platform in early warning and controlling of OPs pollution for safeguarding environmental health and food safety.

Dipyrene-Terminated Oligosilanes Enable Ratiometric Fluorescence Response in Polymers toward Mechano- and Thermo-Stimuli

Developing fluorescent motifs capable of displaying mechano- and thermo-stimuli reversibly and ratiometrically is appealing for monitoring the deformation or temperature to which polymers are subjected. Here, a series of excimer-type chromophores Sin-Py (n = 1-3) consisting of two pyrenes linked with oligosilanes of one to three silicon atoms is developed as the fluorescent motif incorporated in a polymer. The fluorescence of Sin-Py is steered with the linker length where Si2-Py and Si3-Py with disilane and trisilane linkers display prominent excimer emission accompanied by pyrene monomer emission. Covalent incorporation of Si2-Py and Si3-Py in polyurethane gives fluorescent polymers PU-Si2-Py and PU-Si3-Py, respectively, where intramolecular pyrene excimers and corresponding combined emission of excimer and monomer are obtained. Polymer films of PU-Si2-Py and PU-Si3-Py display instant and reversible ratiometric fluorescence change during the uniaxial tensile test. The mechanochromic response arises from the reversible suppression of excimer formation during the mechanically induced separation of the pyrene moieties and relaxation. Furthermore, PU-Si2-Py and PU-Si3-Py show thermochromic response toward temperature, and the inflection point from the ratiometric emission as a function of temperature gives an indication of the glass transition temperature (Tg) of the polymers. The design of the excimer-based mechanophore with oligosilane provides a generally implementable way to develop mechano- and thermo-dual-responsive polymers.

Green synthesis of carbon dots from spent coffee grounds via ball-milling: Application in fluorescent chemosensors

Carbon dots (CDs) are nanoparticles that have gained widespread attention due to their unique properties including multicolored fluorescence, photochemical stability, and biocompatibility. However, the conventional approaches for the preparation of CDs typically require high energy input and toxic reagents. Herein, a sustainable, high throughput approach is reported for producing uniform CDs. Specifically, spent coffee grounds (SCGs) are used to produce coffee ground-derived carbon dots (CFCDs) via a simple wet ball-milling process. The versatility of our approach can be extended to the synthesis of heteroatom-doped CDs and the preparation of CDs from various SCG sources. Also, the functional groups of the CFCDs are manipulated by the addition of citric acids during the ball-milling process to produce carboxylic acid-functionalized carbon dots (CA-CFCDs). CA-CFCDs exhibit highly selective and sensitive detection of Fe3+ ions in aqueous solution via fluorescence quenching, and the relevant quenching mechanism is investigated. Finally, we demonstrate the fluorescent microgel sensor exhibiting a rapid, ratiometric fluorescence change upon the addition of Fe3+ ion by encapsulating the CA-CFCDs within the microgel matrix.