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Shellaiah, M.; Sun, K.W. Carbon Dot-Based Fluorescent Detection of Biothiols. Encyclopedia. Available online: https://encyclopedia.pub/entry/42060 (accessed on 16 January 2025).
Shellaiah M, Sun KW. Carbon Dot-Based Fluorescent Detection of Biothiols. Encyclopedia. Available at: https://encyclopedia.pub/entry/42060. Accessed January 16, 2025.
Shellaiah, Muthaiah, Kien Wen Sun. "Carbon Dot-Based Fluorescent Detection of Biothiols" Encyclopedia, https://encyclopedia.pub/entry/42060 (accessed January 16, 2025).
Shellaiah, M., & Sun, K.W. (2023, March 10). Carbon Dot-Based Fluorescent Detection of Biothiols. In Encyclopedia. https://encyclopedia.pub/entry/42060
Shellaiah, Muthaiah and Kien Wen Sun. "Carbon Dot-Based Fluorescent Detection of Biothiols." Encyclopedia. Web. 10 March, 2023.
Carbon Dot-Based Fluorescent Detection of Biothiols
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This entry is adapted from the peer-reviewed paper 10.3390/bios13030335

Biothiols, such as cysteine (Cys), homocysteine (Hcy), and glutathione (GSH), play a vital role in gene expression, maintaining redox homeostasis, reducing damages caused by free radicals/toxins, etc. Likewise, abnormal levels of biothiols can lead to severe diseases, such as Alzheimer’s disease (AD), neurotoxicity, hair depigmentation, liver/skin damage, etc. To quantify the biothiols in a biological system, numerous low-toxic probes, such as fluorescent quantum dots, emissive organic probes, composited nanomaterials, etc., have been reported with real-time applications.

carbon-dots biothiols detection fluorescence complex-mediated sensors

1. Introduction

Detection and quantification of biologically important species are becoming important for treating infections and diseases existing in living systems [1][2][3]. Therefore, bioimaging of these affected tissues or cells was proposed by using fluorescent organic nanoparticles, inorganic nanostructures, hybrid nanosystems, and composites with authenticated evidence [4][5][6][7][8][9][10][11][12][13]. Among these biologically important species, non-protein biothiols, such as cysteine (Cys; normal blood plasma concentration is between 135 to 300 µM), homocysteine (Hcy; normal blood plasma concentration is between 5 to 15 µM), and glutathione (GSH normal blood plasma concentration is between 1 to 6 µM), play a vital role in many pathological process, clinical disorders, and diseases [14][15][16]. Cysteine plays an important role in protein/peptide synthesis, detoxification, cell metabolism, etc., and lack of cysteine may lead to hair depigmentation, liver damage, skin diseases, and cancer [17][18][19]. On the other hand, elevated cysteine levels can cause neurotoxic disorders [20][21]. Subsequently, homocysteine plays a role quite similar to cysteine. However, elevated concentrations of homocysteine in the blood plasma may lead to hyperhomocysteinemia, which is typically categorized into moderate (concentration = 15–30 µM of Hcy), intermediate (concentration = 30–100 µM of Hcy), and severe (concentration ≥ 100 µM of Hcy) disorders [22][23]. In fact, hyperhomocysteinemia can enhance other disorders, such as osteoporosis, dementia, Alzheimer’s disease, cardiac disorders, etc. [24]. Similarly, deficiency in glutathione decreases immunity and enhances the aging process [25]. Elevated levels of glutathione in the human body may enhance the resistance of cancerous cells to chemotherapy [26]. Individual biothiols play important roles in living systems. For example, they can coordinate with biomarkers to afford cancerous cell bioimaging and predict the therapeutic utilities of numerous drug delivery manuals [27][28]. Thereby, detection and quantification of biothiols is a highly important research topic in this field.
To detect and quantify the biothiols, numerous tactics have been proposed, including colorimetric assay, electrochemical methods, fluorescent imaging, surface enhanced Raman spectroscopy, etc. [29][30][31][32]. Among them, fluorescent imaging is rather impressive in terms of the real-time monitoring of biothiols in living tissues or cells [33][34]. Fluorescent sensing of biothiols can be achieved by using organic probes (undergo a reaction with biothiols), functionalized fluorescent quantum dots, hybrid composite nanomaterials, metal-organic frameworks (MOFs), etc. [35][36][37][38][39][40]. Recently, a smartphone-based surface plasmon-coupled emission (SPCE) platform and photonic crystal-coupled emission (PCCE) technology were also employed in biothiol quantification as well as in biosensing studies [41][42][43][44][45][46]. Among these materials, functionalized fluorescent quantum dots have attracted much attention due to their size, photostability, and unique optical properties (Stokes shifts, wide absorption and optimizable PL, etc.) with respect to surface stabilization [47][48]. The easily synthesizable carbon dots (CDs) with a size of <20 nm, which also belong to the quantum dots category, display exceptional opto-electronic properties and have been applied in energetic applications, sensing, bioimaging, therapy, etc. [49][50][51][52][53]. Numerous reports have discussed the detection ability of CDs towards biothiols in cellular imaging and real samples [54][55][56]. In fact, CDs-based detection of biothiols can be achieved by photoinduced electron transfer (PET), intramolecular charge transfer (ICT), Förster resonance energy transfer (FRET), internal filter effect (IFE), aggregation-caused quenching (ACQ), and aggregation-induced emission (AIE), as demonstrated in published works [48][50]. Similarly, fluorescent CDs-based sensing of biothiols can be performed by observing the “Turn-On” and “Turn-Off” florescent responses via the metal ion–CD pair or CDs-based nanocomposites when exposed to biothiols.
Recently, Khan et al. (2020) delivered a comprehensive review covering reports on both CDs and graphene dots (GQDs)-based biothiols sensing [55]. However, to date, the availability of a review focused on fluorescent CDs-based biothiols detection with information on recent trends, mechanistic aspects, linear ranges, LODs, and real applications is lacking, which allows researchers to deliver this comprehensive review. In this review, the use of emissive CDs in the assay of biothiols (Cys, Hcy, and GSH) is discussed with information on synthesis, photoluminescence quantum yield (PLQY), and demonstrative applications. Moreover, probe/CDs selections, sensory requirements, merits, limitations, and future opportunities for a fluorescent CDs-based biothiols assay are suggested for readers. Figure 1 illustrates schematics of applications and structures of fluorescent CDs-based assay of Cys, Hcy, and GSH.
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Figure 1. Schematic of fluorescent CDs-based assay of Cys, Hcy, and GSH with applications and structures of Cys, Hcy, and GSH.

2. Tactics Involved in CDs Synthesis

Before discussing the CDs-based sensory reports for biothiols, this section briefly describes tactics involved in CDs synthesis. Highly emissive CDs can be synthesized by both (A) top-down approaches and (B) bottom-up approaches. Top-down approaches are categorized into (i) arc discharge, (ii) laser ablation, (iii) chemical oxidation, (iv) electrochemical method, and (v) ultrasonic synthesis. Likewise, the bottom-up approaches can be categorized into (i) microwave synthesis, (ii) hydrothermal method, (iii) solvothermal method, (iv) thermal decomposition, and (v) carbonization/pyrolysis method [57][58].

2.1. Top-Down Approaches

2.1.1. Arc Discharge

With this method, CDs were synthesized by applying a direct-current arc voltage across two graphite electrodes immersed in an inert gas atmosphere. Chao-Mujica et al. reported synthesis of the fluorescent CQDs using this tactic in water [59]. After purification, these CQDs were consumed in cellular imaging studies; therefore, it was noted as a unique top-down approach.

2.1.2. Laser Ablation

With this method, fluorescent CDs were produced by ablating nanosecond pulse laser over a solid carbon target. The as-synthesized CDs were engaged in cellular imaging studies [60]. Doñate-Buendia et al. synthesized the CQDs with a size of 3 nm via laser irradiation in a continuous flow jet and applied them to cellular imaging studies for prolonged periods of time [61]. In fact, this tactic can produce low toxic CDs for numerous bioimaging/biosensing applications [60][61].

2.1.3. Chemical Oxidation

Chemical oxidation, or exfoliation of a disintegrating bulk carbon source, can be achieved by using a strong oxidizing agent, such as H2SO4, HNO3, NaClO3, etc., to produce fluorescent CDs [62]. Desai et al. synthesized fluorescent CDs from muskmelon fruit using sulfuric acid and phosphoric acid as the oxidizing agents [63]. The prepared CDs in the above report were engaged in Hg2+ detection and cellular imaging studies, which have motivated researchers to engage in this synthetic tactic.

2.1.4. Electrochemical Method

In the electrochemical method, the oxidation/carbonization takes place by applying an electric field in a chemical environment to produce the fluorescent CDs [64]. This is a rather straight forward method and has been adopted widely in the production of CDs. Lee et al. synthesized fluorescent CDs by the electrochemical method and employed them in the turn-on recognition of chlortetracycline [65], thereby confirming the effectiveness of this tactic.

2.1.5. Ultrasonic Synthesize

In an ultrasonic process, formation and collapsing of small bubbles in liquid produces a strong hydrodynamic shear force to cut the macroscopic carbon materials into nanoscale CDs [66]. Moreover, CDs with diverse properties can be attained by adjusting the ultrasonic power, reaction time, ratio of carbon sources, solvents, etc. Xu et al. developed the multicolour N-doped CDs from kiwi-fruit juice by the ultrasonic synthesis approach and applied these CDs in investigations of fluorescent inks, sensors, and logic gate operations [67].

2.2. Bottom-Up Approaches

2.2.1. Microwave Synthesis

By irradiating the electromagnetic wave over the sample at a high temperature, CDs can be produced with exceptional PL quantum yield. In fact, electric dipoles in materials are aligned via microwave-assisted excitation. By optimizing precursor and solvent interactions in the microwave synthesis, CDs with hydrophilic, hydrophobic, or amphiphilic properties can be produced for multiple applications [68]. For instance, Liu et al. demonstrated microwave-assisted synthesis of emissive CDs from citric acid, L-cysteine, and dextrin, and employed them in the real-time detection of Cu2+ [69].

2.2.2. Hydrothermal Method

In this method, the reaction mixture in water is placed in a Teflon container and kept in an oven to react hydrothermally at a high pressure and high temperature to produce fluorescent CDs for distinguished applications [57]. For example, Lee et al. synthesized the fluorescent CDs via the hydrothermal method from citric acid, ethylenediamine, and methyl blue and applied them in the “Turn-Off” detection of Hg2+ and ClO− [70].

2.2.3. Solvothermal Method

In contrast to the hydrothermal method, the solvothermal tactic replaces the water with one or more organic solvents. The mixtures are sealed with Teflon and subjected to a steel autoclave under a high temperature and high pressure [71]. This method produces highly fluorescent CDs cost-effectively for various applications. Omer et al. discussed the use of solvothermally prepared phosphorous and nitrogen-doped CDs towards Fe3+ detection [72], and attested the affordability of the tactic.

2.2.4. Thermal Decomposition

Thermal decomposition (via chemical decomposition) by heating the material or compound was engaged in the production of CDs [73]. This tactic is classified as an endothermic process; however, it was rarely used for CDs synthesis due to its complexity. CDs produced from thermal decomposition were also employed in optoelectronic studies. Wan et al. employed the thermal decomposition tactic to synthesize CDs and graphene-like carbon nanosheets and applied them in optoelectronic device fabrication [74].

2.2.5. Carbonization/Pyrolysis

This is a the most cost-effective, facile, and ultrafast method to synthesize CDs. When organic materials are subjected to prolonged pyrolysis in an inert atmosphere, solid residues with a high carbon content or CDs can be produced with a high yield [75]. Esfandiari et al. synthesized fluorescent CDs by pyrolyzing citric acid in different time periods and temperature ranges. The as-synthesized fluorescent CDs employed in cellular imaging studies showed low toxicity, thereby validating the pyrolysis mediated fluorescent CDs synthesis and suggesting feasible drug delivery applications in the future [76].

3. Fluorescence Mechanism, Importance of PLQY, and Desired Size of CDs

3.1. Fluorescence Mechanism of CDs

Synthesized CDs may possess strong, moderate, and weak emission due to the quantum confinement effect, conjugate effect, surface passivation/functionalization effect, surface state, and molecular/carbon-core state properties [77]. Further, the fluorescence of CDs can be tuned via surface passivation, functionalization, doping, and compositing with nanomaterials [78]. In fact, emission of CDs produced by top-down approaches are mostly dependent on surface passivation. On the other hand, bottom-up approaches can produce emissive CDs even without surface passivation [77].

3.2. Importance of PLQY of CDs

The PLQY of CDs defines its capability to convert every absorbed photon into fluorescence emission. The PLQY of CDs serves as a correction factor for the determination of multiparameter fluorescence spectroscopy (MFS) parameters, such as FRET, PL quenching efficacy, incorporation of diverse doping/compositing fluorophores and nanomaterials, complex stoichiometry, and decay profiles, etc. [79]. Further, the use of CDs with a higher PLQY affords the high feasibility of long-term bioimaging and tracking of CDs-based drug delivery systems [80]. To achieve CDs with a high PLQY, surface passivation, functionalization, and doping/compositing with nanomaterials can be used, as stated earlier [77].

3.3. Desired Size of CDs for Biothiols Quantification

Both Top-down and Bottom-up approaches can produce CDs with a size ranging 1–30 nm [81]. The biothiols assay in real water samples can be performed with emissive CDs with a size ranging 1–30 nm [55]. On the other hand, for the detection and quantification of biothiols in intracellular studies, the size of emissive biocompatible CDs must range 1–10 nm [82]. However, in both cases, the lower the size, the greater the emissive properties of CDs to deliver effective analytical results.

4. Representative Mechanism of CDs-Based Fluorescent Biothiols Assay

The CDs-based fluorescent biothiols assay was illustrated by (1) the CD–metal ion pair system [83][84][85][86][87][88][89][90][91][92][93][94][95][96][97][98][99][100][101][102][103][104][105][106][107][108][109][110][111][112][113][114][115][116][117][118][119][120] and (2) CD-nanocomposites [121][122][123][124][125][126][127][128][129][130][131][132][133][134][135][136][137][138][139][140][141][142][143]. Both proposed systems/models deliver the fluorescent response by means of fluorescence recovery or the quenching principle.

4.1. Representative PL Mechanism of CD–Metal Ion Pair in the Biothiols Assay

In general, the interaction of emissive CDs with metal ions (Hg2+, Ag+, Cu2+, Fe3+, and Au3+) led to fluorescent quenching, which recovers due to the effective interaction of biothiols with metal ions, as shown in Figure 2A [87][88][89][90][91][92][93][94][95][96][97][98][99][100][101][102][103][104][105][106][107][108][109][110][111][112][113][114][116][117][118][119][120]. In fact, the metal ions present in the CD–metal ion pair strongly bind with biothiols via an Mn+–S interaction to release emissive CDs and exhibit fluorescence recovery. In contrast, CD–metal ion pair with a certain emission may led to fluorescence quenching upon interaction with biothiols, as seen in Figure 2B; however, it has been reported very rarely [115].
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Figure 2. Representative (A) PL “Turn-On” and (B) PL “Turn-Off” mechanism of the CD–metal ion pair in the biothiols assay.

4.2. Representative PL Mechanism of CD–Nanocomposites in the Biothiols Assay

CD–nanocomposites can be formed by compositing diverse inorganic nanomaterials, conjugation of organic moiety, and doping of metal ions etc., with emissive CDs to afford weakly emissive composites via FRET. These weakly emissive CD–nanocomposites interacts with biothiols to afford two kinds of reaction-based mechanisms, as shown in Figure 3A,B. Biothiols may react with compositing moiety to release emissive CDs (Figure 3A) or interact over the surface of CD–nanocomposites to recovers the fluorescence (Figure 3B). In general, CD–nanocomposites produced by compositing inorganic nanomaterials (such as Au NPs, Ag NPs, and MnO2, etc.) and few organic molecule functionalized composite models [121][122][123][124][125][129][130][131][132][133][134][135][136][137] follows the reaction-based mechanism-2 (Figure 3B). On the other hand, few organic moiety functionalized CD–nanocomposite models [138][139][140] follows the reaction-based mechanism-1 (Figure 3A). Subsequently, CD–DTNB (DTNB = 5,5′-dithiobis-(2-nitrobenzoic acid)) dispersed composite model [141][142] has an initial emission due to IFE, which gets disturbed when it interacts with biothiols, resulting in fluorescence quenching, as visualized in Figure 3C. In fact, biothiols react with DTNB and break it into 2-nitro-5-thiobenzoic acid (TNB), which functionalizes over CDs to afford fluorescent quenching (Figure 3C). A rare report on CD–nanocomposites to afford both PL recovery and quenching for discrimination between biothiols via IFE is available [131]. Therefore, the metal ion doped CD–nanocomposite model for biothiols interactive reaction-based fluorescence quenching was also proposed in Figure 3D. However, to date, only cobalt-doped CDs [143] follows the proposed mechanism.
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Figure 3. Representative (A,B) reaction-based PL “Turn-On” mechanism-1 and mechanism-2 of CD–nanocomposites, (C) PL “Turn-Off” mechanism of CD–DTNB system, and (D) reaction-based PL “Turn-Off” mechanism of metal doped CDs in the biothiols assay.

5. Probe/CDs Selection and Sensory Requirements

The development of fluorescent CDs-based probes towards selective detection and quantification of Cys, Hcy, and GSH must fulfil certain requirements, as illustrated below.
  1. The uniqueness of a CDs-based fluorescent assay of biothiols depends on the size and PLQY. Therefore, to obtain CDs with a proper size and PLQY, it is essential to identify the precursor reactants and suitable synthetic tactics.
  2. To attain high biothiols selectivity in CD–metal complex-mediated detection, the CDs must possess specific selectivity to metal ions with thiophilicity nature (such as Hg2+, Ag+, Cu2+, Fe3+, Au3+, etc.). Therefore, to be able to interact with those metal ions, CDs must possess functional units, such as -NH2 and -COOH, or be doped with N, S, P, etc. However, in the case of doping, the concentration must be carefully tuned to achieve the expected results.
  3. Dye-incorporated CDs towards consecutive ratiometric discrimination of metal ions and biothiols depends on the precise concentration of dye molecules. Thus, it is essential to optimize the dye concentration before designing such innovative probes.
  4. To attain greater sensory responses to biothiols using the CDs incorporated in composites, it is necessary to choose compositing material involved in the detection process/mechanism with thiophilicity.
  5. For dual readout fluorescence and colorimetric detection of biothiols, the CDs must be composited with the colorimetric probe, such as Au NPs. The composition ratio must be fixed to achieve significant results.
  6. Reaction-based sensory responses of CDs to biothiols depend on the reacting units functionalized over the carbon dot surface. Thus, it is necessary to identify molecules to be functionalized over the CD surface at required concentrations that are highly reactive to specific biothiols.
  7. It is essential to categorize the exact mechanisms of the selective sensing of biothiols with CDs-based probes. The coordinative bindings and mechanistic approaches, such as PET, FRET, IFE, and NSET, must be clarified for the emerging new designs.
  8. To commercialize the CDs-based biothiols assay, the exact pH conditions with given details on buffer solutions and concentrations, incubation time, operative temperature, and interference effect must be clarified for researchers.

6. Advantages

CDs-based fluorescent probes and their utilization in biothiols assays have the following advantages, as stated below.
  1. Around 75–80% of reported CDs were synthesized using the one-pot reaction with high PLQY, which is comparable to the current nanoprobes engaged in the detection of different analytes [144][145][146].
  2. CDs-based fluorescent assays of biothiols are equal with inorganic, organic, and hybrid nanoprobes in low toxicity and biocompatibility when the analyte detection is conducted in an aqueous environment [147][148][149].
  3. As discussed in earlier sections, the performance of the CD–metal complex-mediated biothiols detection and quantification is comparable to many metal complex-mediated sensors [150][151][152], which is worthy of attention.
  4. Incorporation of dye molecules and compositing Au NPs with CDs may enhance the ratiometric and colorimetric response towards biothiols. The CDs-based designs are as inspiring as those dye-based sensors [153][154][155].
  5. CDs hold the promise as potentially safe vehicles for biological sample-based biothiols assays due to low in toxicity and high biocompatibility. Moreover, toxicity of CDs can be further reduced by compositing with low toxic nanomaterials, such as Ag NPs, Au NPs, and nanoclusters, to engage in bioimaging and therapeutic applications.
  6. CDs-nanocomposites are comprised of highly selective reactive species (in the presence of biothiols), which can avoid the interference effect. Likewise, CDs are also able to discriminate Cys, Hcy, and GSH via tuning of the pH environment.
  7. Construction of the red-green-blue (RGB) emitting CDs-nanocomposites is possible by mixing CDs with red to blue emissive nanomaterials, which can be utilized for biothiols assay over a broad PL range.
  8. CDs-based fluorometric discrimination of biothiols can be effectively applied in real samples, such as human serum, FBS, plasma, urine, etc. This can be noted as a great advantage towards the development of a unique analytical method.

7. Limitations

CDs-based fluorometric biothiols detection and quantification also have the following limitations, as mentioned below.
  1. Development of fluorescent CDs with high PLQY are limited by the use of precursors and reaction conditions [156][157]. The sensory investigations require careful optimizations of the pH, incubation time, operative temperature, etc.; therefore, real-time detection can be time consuming.
  2. In CD–metal ion pair-based biothiols assays, the formation of metal complexes, such as CD–Hg2+, CD–Ag2+, and CD–Cu2+, may increase toxicity. Hence, the use of such complex-mediated biothiols assays may harm the biological environment or cell lines, which should be carefully examined.
  3. In general, CDs-based specific sensory responses to biothiols are limited by the functional units or doped elements. Thus, careful optimization is mandatory to ensure the existing functional units or doped elements are at required concentrations.
  4. Dye molecules combined with CDs for ratiometric sequential detection of metal ions and biothiols is limited by the concentration and overlapping efficacy of dye molecule, which requires great attention.
  5. CDs-nanocomposites formation for FRET/IFE-based biothiols assays is limited by the composition ratio of CDs and composting material. Otherwise, the primary quenching by FRET or IFE can affected significantly. In case of IFE, it is also essential to clarify the absorbance overlapping of the compositing materials.
  6. The reaction-based biothiols assay is limited by the solid evidence of the mechanistic pathway. In such cases, a model reaction must be conducted to support the proposed mechanism.
  7. Characterization of CDs and detailed mechanistic studies on CDs-based biothiols assay require instruments such as high-resolution transmission electron microscopy (HRTEM), dynamic light scattering (DLS) analyzer, X-ray photoelectron spectroscopy (XPS), fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), etc. Thus, CDs-based biothiols discrimination is limited by the available instruments and cost-effectiveness.
  8. In many reports, CDs-based biothiols quantification was not demonstrated by an interference effect. and discrimination between Cys, Hcy, and GSH was unavailable. Therefore, real sample-based recoveries and bioimaging remain a concern.

8. Conclusions and Perspectives

Fluorescent CDs-based biothiols detection and quantification are discussed in detail. The synthetic methods involved in the fabrication of fluorescent CDs were clearly delivered. Thereafter, the use of (1) a CD–metal ion system and (2) CD–nanocomposites towards biothiols quantification were comprehensively illustrated with their real-time applications in real samples (such as human serum, plasma, and urine) and bioimaging studies. The uniqueness and deficiencies of each report was clearly stated and commented. Finally, the selection of CDs/sensory probe, sensory requirements, merits, and limitations were discussed for the readers. However, some perspective points must be given more attention, as noted below.
  • Many reports of CDs-based fluorescent detection of biothiols followed difficult procedures and did not provide reliable information on the interference effect, which should be rectified to be considered “state-of-the-art”.
  • Up until now, reports on green and red emissive CDs-based assays of biothiols are insufficient, which should be the focus for future research towards a wide range of applications.
  • Although research on using CD–metal ion pairs or composites for detecting Cys, Hcy, and GSH has become the mainstream, not much detail was given on how to distinguish among them. Because Cys, Hcy, and GSH are involved in many different biological processes, this issue should be addressed in the future.
  • In some reports, information regarding the PLQY, exact cause of CDs emission, and PL quenching type (static/dynamic) of CDs with metal ions and during composites formation was not clarified for the readers. These issues should be stated more clearly in future studies.
  • CD–Hg2+ metal complex-mediated fluorescent “Turn-On” sensing of biothiols via a strong Hg–S affinity was proposed in many reports, but none of them were commercialized into practical use [158][159]. This issue should be focused on in the future.
  • The majority of reports delivered recoveries of CD–Hg2+ complex-mediated biothiols assays in biological samples (such as human serum, plasma, urine, etc.) without giving information on the toxicity of the CD–Hg2+ metal complex, which should be clearly addressed in the future.
  • So far, only Hg2+, Ag+, Cu2+, Fe3+, and Au3+ were reported in CD–metal ion pair enabled biothiols assays based on the thiophilicity of metal ions. This approach should be expanded with other thiophilic metal ions, such as Pb2+, Cd2+, Mo4+, etc.
  • Reports on dye molecules incorporated in the CD–metal ion pair system for ratiometric detection of biothiols are still insufficient. Future research should focus on using other dye molecules and justifying the role of dye molecules.
  • The CDs–nanocomposites system for FRET-tuned PL “Turn-On” detection of biothiols can be improved by encouraging more research.
  • To date, only one report is available on the reaction-based PL “Turn-On” dual channel discrimination between Cys, Hcy, and GSH [124], which should be expanded with other biothiols reactive species.
  • Reports on the fabrication of microfluidic paper-based analytical devices from vinyl sulfone clicked CDs for fluorescent assays of biothiols were impressive and could be commercialized. Thus, a similar approach should be strongly encouraged.
  • Only one report is available so far on CD–nanocomposite (Ag NPs/N, S-CDs)-based pH dependence discrimination between Cys, Hcy, and GSH [129], which should be a future research focus.
  • Au NPs and CDs composites displayed dual readout (fluorescent and colorimetric) responses to a specific analyte GSH against Cys and Hcy, which requires more attention in future research.
  • The CD–MnO2 composite system and CD–Br system selectively detects the GSH against Cys and Hcy via redox or specific reactions, thereby such approach can be anticipated for biological applications and towards commercialization.
  • CD–DTNB and Co–CDs (metal doped CDs) composite models showed IFE and reaction-tuned direct recognition of biothiols and Cys, respectively, via the PL “Turn-Off” response against Hcy and GSH. This approach must be improved by more similar research.
  • Reports on CDs-based discrimination of Cys against Hcy and GSH, and GSH against Cys and Hcy are available. However, there is no report on the discrimination of Hcy against Cys and Hcy, which should be the focus towards groundbreaking achievements.
  • The emission of CDs can be enhanced by combining a surface plasmon-coupled emission (SPCE) platform and photonic crystal-coupled emission (PCCE) technology for distinct detection of biothiols at a lower concentration (<nM).
  • Until now, CDs-based fluorescent assays of biothiols lack theoretical support, which should be addressed by density functional theory (DFT) investigations in the future [160][161].
Currently, many research groups are working on developing new CDs-based sensory probes to rectify the aforementioned issues. In terms of PL “On” or “Off” responses to biothiols with real-time applicability, research on CDs-based biothiols assay tactics can be noted as exceptional with great anticipation and excitement.

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Subjects: Biochemistry & Molecular Biology
Contributors MDPI registered users' name will be linked to their SciProfiles pages. To register with us, please refer to https://encyclopedia.pub/register :
Muthaiah Shellaiah
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Kien Wen Sun
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Update Date: 13 Mar 2023
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