Quantum dots for biomedical innovation: overview, applications, and biosafety

1 Introduction

Quantum dots (QDs) are semiconductor-based nanocrystals that are typically composed of group II–VI elements (e.g., CdTe, CdSe, CdS, ZnS, ZnSe, or ZnTe), group III–V elements (e.g., InP or InAs), group I–III–VI elements (e.g., CuInS₂ or AgInS₂), group IV–VI elements (e.g., PbSe, PbS, or PbTe), and group IV elements (e.g., Si, C, or Ge) (Sobhanan et al., 2023). Since their discovery in the 1980s, QDs have garnered significant attention due to their exceptional physical and optical properties (Ekimov et al., 1993). A hallmark feature of QDs is their size-tunable fluorescence: the emission wavelength increases as the particle diameter increases (Figure 1a). This phenomenon, known as the quantum confinement effect, arises from the discrete electronic energy levels in these nanoscale structures. Compared to traditional organic fluorophores, QDs exhibit broad excitation spectra, narrow and symmetric emission bands, remarkable photostability, and high fluorescence brightness (Manikandan et al., 2019; Abdellatif et al., 2022; Panja and Patra, 2023; Zhang J. et al., 2023). The size-tunable emission spectra of CdSe/ZnS QDs are shown in Figure 1b, and a schematic representation of a typical QD probe is provided in Figure 1c. Moreover, their surfaces can be readily modified with hydrophilic ligands, polymers, or biological molecules (e.g., antibodies and peptides), enabling solubility in aqueous buffers and conjugation with targeting moieties for specific biomedical applications (Pham et al., 2021; Han et al., 2020).

Despite these appealing optical and surface properties, the potential toxicity of QDs, particularly those containing heavy metals such as cadmium, remains a subject of intense analysis. QDs’ toxicity is influenced by a multitude of factors, including their intrinsic composition, size, dose, exposure route, and the nature of the surface capping materials (Wang and Tang, 2018). For example, cadmium-based QDs can release toxic Cd²⁺ ions, which cause liver damage primarily by binding to sulfhydryl groups in mitochondrial proteins. This interaction leads to oxidative stress and functional impairment (Zhang Y. et al., 2023; Đukić-Ćosić et al., 2020). Similarly, oxidative stress has also been identified as a key mechanism for Si/SiO₂ QDs. Specifically, Stan et al. (2014) reported that Si/SiO₂ QDs increased ROS generation by 5.5 times (at 200 μg/mL, 48 h) and 7.0 times (at 200 μg/mL, 72 h), respectively, compared with the controls. Furthermore, smaller QDs typically exhibit higher cellular uptake and toxicity than their larger counterparts, with clear dose–effect relationships observed both in vivo and in vitro (Georgeous et al., 2024; Li and Huang, 2018). Surface charge also influences toxicity, as positively charged QDs tend to be more toxic than neutral or negatively charged ones (Chen et al., 2024).

Several mitigation strategies have been developed to address these toxicity concerns. These include coating QDs with protective shells (e.g., ZnS and SiO₂) (Knežević et al., 2013); designing heavy-metal-free alternatives such as cadmium-free QDs (Bechu et al., 2021), metal-doped carbon dots (CDs) (Qi et al., 2024), or carbon quantum dots (CQDs) (Saikia et al., 2023); and applying advanced surface functionalization techniques. Among these, carbon-based QDs have emerged as a “greener” alternative because they contain no heavy metals and are often reported to exhibit minimal or no toxicity in biological systems. For instance, Dhandapani et al. (2022) developed a simple, additive-free, and environmentally friendly method to synthesize high-quality green fluorescent CQDs using fried food waste as a carbon source and deionized water as the solvent. In a separate 2023 study, three types of CQDs derived from biomass waste, such as palm shells, oyster shells, and citric acid, were evaluated across multiple biological models, including HeLa cells, cardiomyocytes, induced pluripotent stem cells, and medaka fish embryos and larvae.

The in vitro assessments revealed no cytotoxicity up to 24 h of incubation (Abu et al., 2023). Further in vivo tests on Japanese medaka fish confirmed the biocompatibility of these CQDs even after 17 days of exposure (Abu et al., 2023). In a similar approach, Saikia et al. (2023) reported the fabrication of biocompatible “green nanoparticles” from waste disposal. Figure 2 illustrates the biocompatible CQDs from petroleum coke and kitchen tea residue via an ultrasonic-assisted oxidation process. These CQDs demonstrated high water solubility, biocompatibility, non-genotoxic behavior, and, in the case of kitchen tea residue-derived CQDs, notable antibacterial activity against various bacterial strains. Collectively, these studies highlight the feasibility of converting biowaste into value-added nanomaterials and underscore the potential of CQDs as promising candidates for biomedical applications, photocatalysis, environmental monitoring, and food industry applications.

This review provided a comprehensive and up-to-date overview of the structure, type, synthesis, property, biomedical application, and biosafety of QDs. Particular focus is placed on the application of QD-based single-virus tracking (QSVT) in studying viral pathogens such as influenza A virus (IAV), Japanese encephalitis virus (JEV), and pseudorabies virus (PRV). By synthesizing the recent advances in the literature, this review aims to clarify the functional roles of QDs, identify current research gaps, and discuss potential strategies for designing safer QDs for use in biomedical contexts.

2 Semiconductor quantum dots

Semiconductor QDs represent a unique class of zero-dimensional nanomaterials that exhibit quantum confinement effects due to their nanoscale dimensions, which are smaller than the exciton Bohr radius (Islam et al., 2025). This quantum confinement leads to discrete electronic energy states, enabling precise tuning of their optical and electronic properties by varying their size and composition. Such characteristics have positioned QDs as revolutionary materials in optoelectronics and biomedical applications. To comprehensively understand QDs, this section systematically explores four critical aspects.

2.1 Types of quantum dots

Quantum dots are typically composed of elements from groups II–VI or III–V of the periodic table. Their emission wavelength is directly proportional to the particle diameter, and both the size and optical properties can be finely controlled through adjustments in the core composition. Based on the elemental constituents, QDs can be categorized into 12 types according to the periodic groups of their constituent elements (Table 1). For example, group I B–VI A QDs such as nitrogen-embedded copper sulfide (Cu₂S) have demonstrated robust anticancer activity and high biocompatibility in breast cancer therapy (Islam et al., 2025). Group II B–VI A QDs, including ZnSe, ZnS, ZnO, CdS, CdSe, CdTe, and HgS, are widely utilized in biomedicine and optoelectronics. An illustrative application is the development of a CdSe/CdS/ZnS-based fluorescence resonance energy transfer (FRET) aptasensor for monitoring Alzheimer’s-related amyloid-β oligomers and tau protein. In this system, dual-color QDs serve as energy donors and Au NRs@PDA serve as acceptors, enabling sensitive detection under single-wavelength excitation (Lu et al., 2022). Group III A–V A QDs such as AlSb, AlAs, AlP, GaSb, GaAs, InAs, InP, and InN have gained attention as promising heavy-metal-free alternatives due to their favorable optical properties and reduced toxicity (Zhang et al., 2017; Zhao et al., 2025). For example, Zhao et al. (2025) developed a dual-modal imaging probe using radioisotope-chelated InP/ZnSe/ZnS QDs combined with copper-64, enabling successful positron emission tomography and fluorescence imaging in tumor-bearing mice. Carbon-based QDs, including graphene quantum dots (GQDs) and CQDs, belong to group IV A and are recognized for their ultra-small size, low toxicity, and excellent biocompatibility. These attributes have spurred considerable interest in biomedical applications (Younis et al., 2020; Peng et al., 2017). Peng et al. (2017) synthesized bone-targeting CQDs with high affinity and specificity for calcified tissues in vivo. Similarly, semiconducting polymer dots (P dots) have emerged as novel near-infrared fluorescent probes. Emitting at approximately 800 nm, P dots exhibit ultrahigh brightness, excellent photostability, and minimal cytotoxicity, making them ideal for deep-tissue bioimaging (Verma et al., 2020; Yuan et al., 2020). Yuan et al. (2020) developed P dots for tracking stem cells and cancer cells in vivo, achieving a labeling intensity that was approximately 10,000 times higher than that of unlabeled controls. Looking forward, next-generation smart biomaterials such as 0D titanium carbide (Ti₃C₂) and Nb₂C MXene QDs have shown intrinsic immunomodulatory properties, including the selective suppression of human lymphocyte activation, thereby supporting material-assisted post-injury tissue regeneration (Rafieerad et al., 2019).

TABLE 1. Classification of QDs according to their chemical composition.

2.2 General properties of quantum dots

QDs exhibit a suite of exceptional photophysical properties that make them highly appealing for a wide range of applications. These include high brightness, remarkable photostability, broad excitation spectrum, size-tunable emission, and large significant Stokes shift. These optical characteristics stem from the fundamental physical properties of QDs, particularly their nanoscale dimensions, which lead to a markedly increased surface-to-volume ratio and a significant proportion of surface atoms relative to the bulk material. This high surface-to-volume ratio profoundly influences the dynamics of photo-generated charge carriers, often enhancing or reducing their transfer rates due to the high density of surface states.

The optical properties of QDs, such as optical absorption (measured via photoluminescence excitation, PLE), quantum yield, luminescence intensity, spectral profile, and aging behavior, are intimately tied to their surface states, which are, in turn, determined by their size, shape, and surface structure (Weiss, 2013). A key manifestation of this structure–property relationship is the quantum confinement effect, which becomes significant when the QD diameter is comparable to the de Broglie wavelength of the charge carriers. This effect is primarily responsible for the size-dependent tuning of the absorption and emission spectra.

Quantitatively, the emission wavelength of QDs is directly determined by their bandgap energy (E _*g*), which is strongly dependent on the particle size due to quantum confinement (Oglah et al., 2025). When the QD radius (R) is smaller than the exciton Bohr radius, electrons and holes are physically confined, leading to an increase in their kinetic energy and, thus, a broadening of the effective bandgap. Brus (Chukwuocha et al., 2012) presented the first theoretical calculation for semiconductor nanoparticles (NPs) (using CdS and CdSe as examples) based on “effective mass approximation” (EMA). In this approximation, as shown in Equation 1, an exciton is considered to be confined to a spherical volume of the crystallite, and the mass of the electron and hole is replaced with effective masses (me and mh) to define the wave function (Brus, 1984).where

E g q d = E b u l k + h 2 8 R 2 1 m e * + 1 m h * ‐ 1.786 e 2 4 π ε 0 ε r R 2 , (1)

E _g(qd) = band gap energy of quantum dot;

E_bulk = band gap energy of the bulk semiconductor;

R = radius of quantum dot;

m e * = effective mass of excited electron;

m h * = effective mass of excited hole;

h = Planck’s constant.

Kayanuma (1988) accounted for the electron–hole spatial correlation effect and modified the Brus equation. Based on the modified equation, as shown in Equation 2, the size-dependence on the band gap energy of quantum dots can be quantified as follows:where

E g q d = E b u l k + h 2 8 R 2 1 m e * + 1 m e * ‐ 1.786 e 2 4 π ε 0 ε r R 2 ‐ 0.248 E * Ry , (2)

h = Planck’s constant

m e * = effective mass of excited electron;

m h * = effective mass of excited hole;

R = radius of quantum dot;

e = electronic charge

In summary, in QDs, excitons are under a three-dimensional confinement that leads to the quantization of the electron and hole energy levels. Semiconductor materials sized less than double the bulk exciton Bohr radius undergo quantum confinement. The quantum confinement effect will be observed in CdS, CdSe, and CdTe NMs when their particle diameters are less than ∼5.8 nm, 11.2 nm, and 20.0 nm, respectively. Thus, the bulk exciton Bohr radius and, by extension, the composition determine the size from which a material exhibits quantum confinement effects and, consequently, the unique optical and electronic properties of QDs. Since electrons within QDs are confined to discrete energy levels, the wavelengths associated with the formation and recombination of excitons (i.e., the UV–Vis absorption and fluorescence emission maximum wavelengths of QDs, respectively) are tunable by changing the particle’s size. With decreasing size, the wavelengths of fluorescence and absorption decrease. For example, CdSe QDs exhibit emission wavelengths ranging from ∼500 nm (green) to ∼650 nm (red) as their diameter increases from 2 nm to 6 nm.

The correlation between the QD size and their electronic structure is well-illustrated by GQDs. Peng et al. (2012) discovered that the band gap of GQDs and, consequently, their photoluminescence (PL) emission can be systematically varied by controlling their size. In their study, GQDs synthesized at different temperatures (80 °C, 100 °C, and 120 °C) exhibited a distinct blue shift in absorption from 330 to 270 nm as the synthesis temperature increased (Figures 3a,b). Furthermore, the optical band gap decreased from 3.90 to 2.89 eV with increasing GQD size (Figures 3c,d), confirming that the energy gap can be precisely engineered through dimensional control.

Beyond size control, the architectural design of QDs, specifically the formation of a core/shell structure, profoundly influences their electronic structure and optical performance. Due to their high surface-to-volume ratio, QDs possess a high density of surface defects/traps, which severely reduce the photoluminescence quantum yield (QY) and promote blinking. An effective strategy to mitigate these issues is surface passivation through the growth of an outer shell of a different material or composition, leading to so-called core/shell QDs. Such a shell reduces the surface defects and modifies the confinement potential, thereby broadening the absorption spectrum, accelerating charge carrier transfer, and suppressing exciton-recombination losses. Consequently, core/shell QDs exhibit markedly improved QY and reduced blinking. For instance, Kim et al. (2022) recently reported alloy-typed CdSeZnS/ZnS core/shell QDs with an initial QY as high as 98.0%, which remained at 84.7% even after hydrophilic surface modification—a decrease of only ∼13.6%. In contrast, conventional multilayer QDs lost more than 50% of their QY under the same treatment. The superior performance of core/shell QDs underscores the critical role of shell engineering in minimizing surface defects, enhancing photostability, and maintaining high QY for demanding bioimaging applications. Therefore, achieving precise control during synthesis is paramount, as the intrinsic properties of QDs are governed by multiple factors, including the size, shape, defect density, impurities, and crystallinity. The quantum confinement effect is a dominant factor for the monotonic shift in absorption with changes in size. However, non-monotonic behaviors in the photoluminescence spectra have also been observed, such as when QDs’ size increases to approximately 17 nm. Such complex spectral patterns can often be attributed to concomitant changes in the QDs’ shape and edge structure, which occur during growth, highlighting that size is not the sole determinant of their optical behavior.

In core-only QDs, the large surface-to-volume ratio leads to abundant dangling bonds and lattice defects, which act as trap states for charge carriers, thus severely reducing the photoluminescence quantum yield and promoting blinking. The growth of a wider-bandgap semiconductor shell (e.g., ZnS on CdSe) passivates these surface defects, eliminates dangling bonds, and spatially confines excitons away from the surface, thereby reducing non-radiative recombination. As a result, the core/shell QDs exhibit markedly improved QY and suppressed blinking. For instance, Kim et al. recently reported alloy-typed CdSeZnS/ZnS core/shell QDs with an initial QY as high as 98.0%, which remained at 84.7% even after hydrophilic surface modification—a decrease of only ∼13.6%, whereas conventional multilayer QDs lost more than 50% of their QY under the same treatment. The superior performance of core/shell QDs underscores the critical role of shell engineering in minimizing the surface defects, enhancing photostability, and maintaining high QY for demanding bioimaging applications. Therefore, understanding the electronic structural differences between core-only and core/shell QDs is essential for designing high-performance quantum dots for optical and biomedical applications.

2.3 Challenge of quantum dots synthesis

An array of physical, chemical, and biological methods have been developed to synthesize different QDs, and the methods are mainly divided into two important methodologies, namely, the top-down or bottom-up approaches (Chan et al., 2002). Top-down methods involve the fragmentation of larger precursor materials into nanoscale QDs using techniques such as molecular beam epitaxy, ion implantation, and electron-beam lithography (Mattoussi et al., 2012). In contrast, bottom-up approaches rely on the self-assembly of smaller molecular precursors into larger QD structures. Given that the optical properties that are most relevant to bioimaging are predominantly exhibited by colloidal QDs that are typically synthesized via bottom-up routes, this section will focus exclusively on these methods, including colloidal synthesis, non-injection synthesis, electrochemical synthesis, bio-template mediated synthesis, and biogenic synthesis (Figure 4).

2.3.1 Colloidal synthesis: precision at the laboratory scale

Colloidal synthesis, called organometallic synthesis, produces QDs from metal-organic or inorganic substances and disperses them in solvents (Pu et al., 2018). In this process, semiconductor precursors are rapidly injected into a hot solvent containing coordinating ligands, leading to a burst of nucleation followed by controlled growth (Figure 4a). The classic synthesis of high-quality CdSe QDs using Cd(CH₃)₂ and elemental Se established a model system (Murray et al., 1993). However, the toxicity and instability of such precursors prompted the development of greener alternatives. Peng and Peng (2001) successfully replaced Cd(CH₃)₂ with the safer CdO, while subsequent innovations introduced less hazardous solvents such as octadecene (Yu and Peng, 2002) and paraffin liquid (Deng et al., 2005), demonstrating that high optical quality could be maintained through precise optimization of the reaction parameters (e.g., injection speed and temperature). Despite its unparalleled ability to produce monodisperse and high-quality QDs, the hot-injection method faces challenges in scalability and reproducibility for core/shell structures, which limits its practicality for large-scale biomedical applications.

2.3.2 Non-injection synthesis: scalable and greener synthetic routes

Non-injection organometallic synthesis was developed to address the scalability limitations of hot-injection methods (Figure 4b). This one-pot approach involves heating all the reagents together, enabling the large-scale production of QDs. For instance, Park et al. (2013) synthesized Gram quantities of zinc blende CdSe QDs using various Cd–fatty acid salts, such as Cd-myristate (C14), Cd-palmitate (C16), and Cd-stearate (C18). Further advancing this route, Zhang et al. (2012) developed an air-stable and scalable method for producing high-quality core/shell QDs. Ding et al. (2017) pushed the boundaries [Cd(CH₃COO)₂·2H₂O, Na₂S·9H₂O, oleic acid, and oleylamine] further by establishing a low-temperature, ambient-air synthetic route for doped CdS QDs with tunable emissions, demonstrating their direct application in high-color-rendering-index white LEDs. This progression highlights a clear trend toward more industrially viable and environmentally benign synthesis.

2.3.3 Electrochemical and bio-template synthesis

Electrochemical synthesis represents a green alternative that eliminates the need for external chemical reducing agents (Figure 4c). This method allows for precise control over the generation of reactive species, leading to reproducible and clean QD synthesis. Ribeiro et al. (2013) and Freitas et al. (2017) utilized this approach to synthesize various CdX (X = Te, Se, or S) QDs in aqueous solutions. Valappil et al. (2017) extended its utility by electrochemically converting bulk WS₂ directly into luminescent QDs at room temperature. The prepared WS2 QDs had an average diameter of 3 ± 1 nm, showed a higher photoluminescence (PL) quantum efficiency (5%), and exhibited an excitation wavelength-dependent photoluminescence. This electrochemical transformation was expected to have a greater impact on the design and development of nanostructures derived from 2D materials.

Bio-template-mediated synthesis is a technique that uses biological molecules (e.g., proteins, DNA, viral particles, and bacteria) as templates to guide the formation of QDs (Figure 4d). Biological components are utilized in this process to improve the stability and biocompatibility of quantum dots while also improving their performance. Nithyaja et al. (2012) showed the choice of two biological templates (DNA and BSA) that significantly influence the final optical properties of the CdS nanoparticles. Ai et al. (2024), Ai et al. (2022) demonstrated the assembly of PbS QDs on proteinaceous templates, enhancing their mechanical and photoluminescent properties. The prepared QDs demonstrated improved modulus and hardness, superior hydrophilic reinforcement, and excellent PL properties. Similarly, DNA-templated CdS nanoparticles exhibited low cytotoxicity and potential as biocompatible probes for imaging and therapy (Reena et al., 2024). In this method, uniform voids in porous materials serve as hosts, and the DNA template was used as smart glue for assembling in nanoscale. The prepared CdS nanoparticles exhibited visible fluorescence and low cytotoxicity in HeLa cells. This biomimetic approach offers exquisite control over the size, morphology, and optical properties of the produced QDs.

2.3.4 Biogenic synthesis: nature-inspired biocompatibility

Biogenic synthesis is a novel biotechnology-based approach that utilizes living organisms (bacteria, fungi, and plants) or their components to produce QDs (Figure 4e). This eco-friendly approach often yields QDs with superior biocompatibility. Mi et al. (2011) utilized engineered Escherichia coli that expressed a specific CdS-binding peptide to synthesize CdS QDs. The QDs, which were subsequently isolated through cell lysis and purification, demonstrated high biocompatibility, making them promising probes for bio-labeling and imaging applications. Further demonstrating the versatility of biogenic synthesis, Borovaya et al. (2015) utilized the fungus Pleurotus ostreatus as a platform to for the eco-friendly production of CdS QDs. By incubating P. ostreatus mycelium with inorganic cadmium sulfate and sodium sulfide, CdS QDs sized 4 nm–5 nm were synthesized. In a complementary approach, Raja and Sundaramurthy (2021) utilized betel leaves (Piper betle) in a hydrothermal process to synthesize fluorescence CQDs, which served as a selective sensor for Fe³⁺ (detection range: 50 nM–150 nM) in the presence of competing metal ions. While this field is still developing, it holds significant potential for generating QDs with minimal environmental impact and native biological recognition capabilities.

2.4 Surface functionalization of quantum dots

Although QDs demonstrated impressive performance, various application scenarios—particularly in biomedicine—impose more specific requirements on their properties. Consequently, it is essential to make targeted adjustments and modifications to the chemical structure of QDs to meet these specific application needs. The synthetic modification of QDs primarily focuses on surface functionalization and element doping. In the context of surface functionalization, three key parameters must be considered, namely, surface functional groups, surface charge, and ligand exchange dynamics. Optimization of these parameters can effectively improve their dispersibility and stability in different solvents and modulate their biological interactions. This section provides a systematic discussion of these three parameters and their roles in tailoring the performance of QDs for biomedical applications.

2.4.1 Surface functional groups

Surface functional groups directly determine the physicochemical interaction of QDs with their surrounding environment. Common functional groups, including carboxyl (-COOH), amine (-NH₂), thiol (-SH), and hydroxyl (-OH) groups, along with organic ligands such as oleylamine (OAm) and oleic acid (OA), are introduced via ligand exchange or coating strategies. In biological environments, surface modifications can mitigate protein corona formation and modulate cellular uptake issues, thereby improving the biocompatibility and targeting efficiency of QDs. Li L. et al. (2020) prepared three kinds of InP/ZnS QDs with -COOH, -NH₂, and -OH surface groups and compared the biodistribution in vivo. The results demonstrated rapid uptake in the liver and spleen without significant aggregation, indicating that the surface chemistry influences organ distribution while maintaining colloidal stability. Dumbgen et al. (2023) synthesized OAm- and chloride ligands-stabilized InP QDs though amine–halide co-passivation with diethylamino phosphine, allowing subsequent exchange with carboxylates or thiolates to tailor the surface chemistry. This approach enables precise control over surface functionality, which is essential for optimizing the performance of QD in biological media. Beyond passivation, surface functionalization also provides anchoring sites for secondary coatings and targeting ligands. For instance, a ZnS shell not only passivates surface defects and enhances the photoluminescence quantum yield but also increases water solubility and offers abundant sites for further functionalization, making such core/shell QDs suitable for bioimaging and cancer therapy. In a representative study, PEG and folate molecules were successfully conjugated to CdTe@SiO₂ particles through the amino groups in the water phase (Lu et al., 2014). The prepared folate–PEG-functionalized QDs exhibited improved hydrophilicity and reduced toxicity, holding promise for tumor imaging in the near future. Expanding on this strategy, Zhao et al. (2025) developed a novel PET/fluorescence imaging probe based on InP/ZnSe/ZnS QDs, wherein the ZnS QDs were functionalized with polyethylene glycol terminated with either a methoxy group or a DOTA chelator group. The dual-modal probe enabled multi-scale imaging from the whole body to the subcellular levels, contributing to more precise tumor diagnosis. Therefore, rational design of surface functionalization strategies is essential for developing QD-based nanoplatforms with predictable and desirable biological interactions.

2.4.2 Surface charge

Surface charge plays a more dominant role than protein corona formation in determining the cytotoxicity, cellular uptake, and biodistribution of QDs (Barbalinardo et al., 2025; Fröhlich, 2012). Positively charged QDs generally exhibit enhanced efficacy in imaging, gene transfer, and drug delivery; however, they are also associated with higher cytotoxicity and hemolytic activity. In a systematic study, Liu et al. (2015) compared three commercially available QDs with different surface coatings to assess the influence of the surface charge on cellular uptake, cytotoxicity, and in vivo biodistribution. Charged QDs (both positive and negative) entered cancer cells and macrophages more efficiently than neutral QDs, with negatively charged QDs showing the highest internalization. Notably, positively charged QDs—but not negatively charged ones—exhibited severe cytotoxicity, which was attributed to the disruption of cell membrane integrity and induction of reactive oxygen species production. Negatively charged QDs generally show reduced non-specific uptake and better biocompatibility, although extreme negative charge can induce protein denaturation and complement activation (Barbalinardo et al., 2025). Similarly, Zhang et al. (2025) prepared a series of drug-loaded liposomes with different surface charges and found that negatively charged liposomes exhibited better mucus penetration and good biocompatibility (low cytotoxicity, better blood compatibility, and mild tissue inflammation), but with poor cellular uptake by immune cells. These findings indicate that liposomes with a negative charge (−1 to −3 mV) may serve as promising inhaled drug carriers that balance the efficacy and biocompatibility. Furthermore, surface charge modulates the composition of the protein corona, which, in turn, influences QD uptake and macrophage viability. Liu Y.-Y. et al. (2025) analyzed the effects of surface charge on the uptake, intracellular distribution, and exocytosis of CdSe/ZnS QDs in Raw264.7 macrophages. Expanding on this, Arezki et al. (2022) performed the first quantitative proteomic analysis of the protein corona formed on CDs with different surface charges. CDs with high ζ-potential (>+30 mV) and charge density (>2 μmol mg⁻¹) showed the highest internalized value, which was strongly correlated with a corona enriched in vitronectin, fibulin, fetuin, adiponectin, and alpha-glycoprotein. Collectively, these findings underscore that the surface charge is a critical determinant of the biological interactions of QDs, directly influencing protein corona composition, cellular uptake pathways, toxicity profiles, and in vivo fate.

2.4.3 Ligand exchange dynamics

Ligand exchange dynamics, also referred to as cap exchange, occur during the substitution of native hydrophobic ligands with hydrophilic ones via a mass-action-driven process (Nabil and Megahed, 2024). In this process, the original hydrophobic ligands are displaced by water-soluble bifunctional molecules, wherein one functional group anchors to the QD surface (e.g., thiols binding to the ZnS shell), while the other group—typically -OH, -COOH, or -NH₂—imparts hydrophilicity and provides a reactive handle for subsequent conjugation with biomolecules such as antibodies, proteins, or drugs. This bifunctional design markedly improves the aqueous dispersibility, colloidal stability, and biocompatibility, as evidenced by functionalized silicon carbide nanostructures that maintain structural integrity in physiological environments. For instance, Saad et al. (2026) systematically evaluated silicone quantum dots functionalized with -COOH and -OH groups and demonstrated that dual-surface carboxylation serves as an optimal strategy for designing high-performance, reversible, and targeted silicon-based drug delivery systems. Notably, dual functionalization with -COOH significantly enhanced the adsorption of the anticancer drug thioguanine, achieving the highest binding energy (−2.04 eV) and the shortest interaction distance (2.54 Å). More importantly, the ligand exchange dynamics directly govern the biological identity of QDs: rapid or incomplete ligand exchange can lead to the loss of surface coating, exposure of hydrophobic cores, and uncontrolled protein corona formation, thereby altering the cellular uptake pathways and triggering non-specific immune recognition. Therefore, characterizing and controlling ligand exchange kinetics is essential for the rational design of QDs with predictable nano–bio interfacial behavior.

2.5 Characterization of QDs

The properties and performance of QDs are intrinsically governed by their structural, optical, and surface characteristics. This section outlines the principal analytical techniques utilized to assess QDs across three key parts, namely, structural and morphological features, size-dependent optical behavior, and surface chemistry. Together, a systematic and multi-faceted characterization approach is essential to establish clear structure–property relationships and guide their rational design for specific applications.

2.5.1 Structural and morphological analysis

The size and size distribution of QDs, which dictate their electronic structure and emission wavelength via the quantum confinement effect, are initially estimated using dynamic light scattering (DLS) (Xing et al., 2019), which provides the hydrodynamic diameterMulti-angle dynamic light scattering (MADLS) is a new method for providing better resolution for multi-modal populations and more accurate particle size distributions than traditional DLS (Coones et al., 2025). For precise determination of the core size, morphology, crystallinity, and lattice structure, high-resolution microscopy techniques are indispensable. Scanning electron microscopy (SEM) (Torchynska et al., 2007), transmission electron microscopy (TEM) (Gnanasekaran et al., 2015), and atomic force microscopy (AFM) (Cockins et al., 2010) are widely conducted to provide direct visualization of QDs at the nanoscale, while techniques such as powder X-ray diffraction (XRD) provide complementary data on the crystal phase and quality (Holder and Schaak, 2019).

2.5.2 Optical property analysis

The size-tunable optical properties of QDs are quantitatively assessed using spectroscopic methods. Optical characterization of QDs is usually provided by UV–Vis and PL, which offer fast, non-destructive, and contactless option. UV–Vis absorption spectroscopy determines the absorption onset and band gap, while PL spectroscopy measures the emission maxima, spectral shape, and quantum yield and (Hosseinpour-Mashkani and Sobhani-Nasab, 2017). Time-resolved PL spectroscopy further probes excited-state dynamics and surface defect states, offering insights into non-radiative recombination pathways (Van Sark et al., 2002).

2.5.3 Surface chemistry and functionalization analysis

Given the high surface-to-volume ratio of QDs, their colloidal stability, biocompatibility, and functional performance are critically dependent on the surface chemistry. This makes the characterization and modifications of surface ligands paramount. For QDs intended for biomedical applications, such as bioimaging and intracellular tracking, QDs have been coated with different surface ligands such as glutathione (Merkoçi et al., 2007), amphiphilic polymers (Gallagher et al., 2018), and N-acetyl-L-cysteine (Barriera and Bailón-Ruiz, 2020) to enhance the water solubility and stability and reduce toxicity. Overall, a combination of sophisticated techniques is utilized to validate and optimize these surface designs. Nuclear magnetic resonance (NMR) (Morris-Cohen et al., 2012) and Fourier transform-infrared spectrophotometry (FT-IR) (Cooper et al., 2011) are effective in confirming the successful binding and identity of organic ligands. X-ray photoelectron spectroscopy (XPS) is particularly powerful for determining the elemental composition and chemical states at the QDs’ surface, providing direct evidence of successful conjugation or shell formation, and it has been the technique of choice for the analysis of the surface of a variety of QDs (Virieux et al., 2012; Rath et al., 2008). Size exclusion chromatography (SEC) (Wu et al., 2016) is valuable for assessing hydrodynamic size changes and purity following surface functionalization. The insights gained from these techniques—such as verifying ligand density, assessing shell coverage, or identifying surface oxidation—directly inform the iterative refinement of synthesis and coating protocols. For instance, XPS data revealing incomplete surface passivation can guide adjustments in shell growth conditions to minimize surface traps and enhance photoluminescence quantum yield, which is a critical parameter for sensitive imaging (Isaacs et al., 2021).

3 Application of quantum dots in biomedical field

QDs technology emerged in the late 1970s, with early research primarily concentrated in the field of optoelectronics. A pivotal shift occurred in 1998 when QDs were first successfully utilized as biological probes in cellular systems, marking the inception of their application in biomedicine (Chan and Nie, 1998; Bruchez et al., 1998). A typical QDs platform features core/shell–ligand architecture. As shown in Figure 5, the semiconductor core indicates the fundamental optical properties, while an inorganic shell, often composed of a wider bandgap material, encapsulates the core to enhance its photostability and quantum yield. The outer layer provides aqueous solubility, biocompatibility, and further functional versatility. This layer can be conjugated with various biomolecules, such as peptides, antibodies, or oligonucleotides, for targeted recognition or modified with polymers such as PEG to improve pharmacokinetics (Liu et al., 2015). Collectively, these advantageous properties, including tunable emission, high brightness, and surface tailoring ability, establish QDs as highly promising nanomaterials for a wide range of biomedical applications. To provide a clear comparative overview of how different QD types perform across key biomedical fields, Table 2 summarizes the representative studies in cellular imaging, drug delivery, and single-virus tracking, highlighting their core structures, surface modifications, and critical performance metrics. The following Sections (3.1–3.3) discuss these applications in detail.

TABLE 2. Comparison of different types of quantum dots in cellular imaging, drug delivery, and single-virus tracking.

3.1 Imaging cells

QDs have emerged as powerful fluorescent probes for cellular imaging, largely due to their superior photostability and high brightness, which surpass the performance of conventional organic dyes and significantly reduce interference from cellular autofluorescence. These attributes make QDs particularly suitable for immunolabeling applications. A common strategy involves conjugating QDs to specific antibodies or utilizing biotin–streptavidin affinity systems to label cell-surface antigens. Fountaine et al. (2006) demonstrated the utility of multiplexed staining using streptavidin-conjugated QDs of distinct emissions, enabling simultaneous detection of seven specific antigens on tissue specimens. This approach facilitated detailed analysis of the spatial organization of biomarkers and compartmentalization of tissues with improved sensitivity, though it carried a risk of non-specific signal transfer via the streptavidin-QDs. This limitation can be mitigated by using pre-conjugated QD–antibody complexes for one-step staining, which also shortens the assay time, as exemplified by Sweeney et al. (2008) in a three-antibody multiplex stain on tonsillar tissue. More recently, QDs-based multiplexed imaging and spectral analysis have been reported to dissect the complex composition of tumor stroma, providing novel insights into tumor invasion and informing anti-cancer strategies (Peng et al., 2011; Hu et al., 2014; Chen et al., 2022).

A critical application of QDs imaging lies in cancer theranostics, where tracking cancer cells can guide therapy and monitor early treatment response or drug resistance (Kunachowicz et al., 2024). Conventional organic dyes are unsuited for deep-tissue imaging due to photobleaching and strong tissue absorption of visible light (Jacques, 2013; Clarke et al., 2010). In contrast, QDs-based probes offer a robust alternative. A step toward advancement involves conjugating QDs with uniformly oriented, high-affinity, single-domain antibodies (sdAbs or nanobodies) to form tumor-targeting nanoprobes (e.g., sdAb-HER2-QDs and sdAb-CEA-QDs) (Sukhanova et al., 2021). After quality control, sdAb-HER2-QDs/sdAb-CEA-QDs nanoprobes were used for deep-tissue imaging in animal models, successfully detecting micro-metastases and disseminated tumor cells in HER2- or CEA-expressing cancers. Confocal and two-photon microscopy confirmed specific binding to target cells (e.g., BT474 and MDA-MB231 breast cancer lines), visualized tumor invasion in the intestinal crypts and liver, and identified metastases in the lymph nodes and liver. These findings proved the potential of engineered QDs probes for tracking and treating cancers through high-resolution deep-tissue imaging.

Despite their advantages, QDs present certain limitations in cellular imaging concerning photobleaching, fluorescence stability, and the signal-to-noise ratio. While significantly more photostable than organic dyes, QDs are not entirely immune to photobleaching, especially when QDs are subjected to higher energy light, their photo brightening and photodarkening conjointly determine the photostability, which can degrade the ligand/shell structure or surface photo-oxidation (Chen G. H. et al., 2023). The fluorescence stability of QDs also depends on the surface chemistry (Bilan et al., 2015), positive sorption behaviors in the ligand exchanges (Jin et al., 2021), aggregation in various biochemical buffers (Boldt et al., 2006), and interaction with biological systems (Le et al., 2022), all of which can lead to quenching or blinking, thereby compromising quantitative imaging. Blinking, the intermittent fluorescence intermittency, arises from charge trapping at the surface defect sites or in the core, where photoexcited electrons or holes are temporarily immobilized, leading to non-radiative recombination. This phenomenon severely affects single-molecule tracking and quantitative imaging reliability. Three main mechanisms have been proposed to explain blinking, namely, Auger blinking due to QD charging, band-edge carrier (BC) blinking due to band-edge carrier trapping, and hot carrier (HC) blinking due to hot carrier trapping (Yang et al., 2025). Several effective strategies have been developed to suppress blinking, including surface modifications, reduction of Auger recombination, introduction of extra electron transfer channels, all-optical control, and enhancement of radiative recombination (Ahmad et al., 2025; Ghosh et al., 2021; Chouhan et al., 2021; Xie et al., 2022; Thomas et al., 2018; Shi et al., 2021). Notably, through advanced shell engineering, nearly non-blinking QDs with an on-state fraction exceeding 99% have been achieved through advanced shell engineering, accompanied by near-unity photoluminescence quantum yield (Ahmad et al., 2025). These advancements markedly improve the performance of QD-based devices and are particularly beneficial for biomedical applications such as single-particle tracking, in vivo and in vitro imaging, and biosensing. Furthermore, while their brightness improves the signal-to-noise ratio relative to autofluorescence, comparatively large hydrodynamic diameters of QDs can increase non-specific binding, affect cell membrane adsorption and internalization (Dong and Irudayaraj, 2012), and even hinder efficient tissue penetration (Zhang et al., 2018). Ongoing efforts to engineer smaller, brighter, and more biostable QDs, along with improved surface passivation and targeting strategies, are focused on overcoming these challenges to fully realize their potential in sensitive and quantitative bioimaging.

3.2 Drug delivery

In recent years, NPs such as micelles, liposomes, gold colloids, and polymers have gained significant traction in drug delivery research. However, a common limitation of many conventional NPs is the lack of real-time high-resolution imaging signals after in vivo administration. In contrast, QDs have multiple advantageous features as drug delivery systems, including their ease of fabrication, capability of conjugation to a wide variety of drugs, tuned physico-chemical properties, and interesting optical properties making them traceable drug carriers that can easily be monitored after administration.

Yezhelyev et al. (2008) reported a siRNA delivery and imaging system by coating QDs with a proton-sponge polymer. This platform enabled real-time dual-modality tracking of the entire delivery process—from cellular penetration and endosomal release to carrier unpacking and intracellular siRNA localization. However, endosomal escape is a critical bottleneck for QDs-based intracellular delivery. Following endocytosis, most nanocarriers are trapped in endo-lysosomal compartments, where hydrolytic enzymes degrade therapeutic drugs (Wang X. et al., 2025). The proton-sponge effect, utilized by polymers such as PEI and poly(beta-amino esters), induces osmotic swelling, leading to endosomal rupture (Qiu et al., 2023). Recent molecular dynamics simulations by Steinegger et al. (2026) further revealed that hydrophilic polymers such as PEI primarily utilize electrostatically driven membrane attachment and osmotic stress, whereas amphiphilic polymers (e.g., PBAE with 70% oleylamine) induce membrane fusion and lipid extraction, achieving higher escape efficiency at the cost of increased cytotoxicity. Lipid nanoparticles escape via membrane fusion, forming small pores with minimal endosomal damage and inflammation (Zhao et al., 2026). Photochemical internalization using light-triggered ROS generation offers another orthogonal strategy (Liu M. et al., 2025; Huang et al., 2025). Benefiting from the above escape mechanisms (especially the proton-sponge effect), the QD–siRNA system developed by Yezhelyev et al. achieved efficient cytosolic delivery. Compared with the transfection agent, the gene-silencing efficiency of the QD–siRNA nanoparticles achieved a 10–20-fold improvement. .

Building on the importance of endosomal escape, the intracellular trafficking pathways that determine the fate of QDs-based nanocarriers must also be carefully considered. Similarly, polymeric micelles have been engineered as integrated theranostic vehicles (Bagalkot et al., 2007; Chakravarthy et al., 2011). Kumar et al. (2012) exploited the hydrophobic core of the DSPE-PEG-based micelles for encapsulation of the anti-cancer drug doxorubicin (Dox) and CdSe QDs together to create a new drug-delivery vehicle, where the QDs are for diagnosis and Dox is for therapy. QDs micelles showed >95% cell survival after a 24-h incubation time period, demonstrating biocompatibility of the QD micelles. As shown in Figure 6, QDs–Dox micelles at a Dox concentration of 5 μg/mL showed a strong cytotoxic effect (∼50% cell survival), while at the same concentration, free Dox showed more than 65% cell survival, depicting the efficient delivery of Dox to the HeLa cells. Confocal microscopy confirmed time-dependent cellular uptake, validating the potential of such QD-loaded micelles for image-guided chemotherapy. The cellular internalization of these micelles is governed by their size, surface charge, and trafficking (Xiao et al., 2011). Generally, QDs with smaller diameters and neutral or negative surface charge enter cells predominantly via clathrin-mediated endocytosis, which delivers cargo to early endosomes and subsequently to lysosomes (Damian‐Buda and Boccaccini, 2026). In contrast, larger QDs or surface modifications such as using folic acid, cholesterol, or transferrin may engage caveolin-mediated endocytosis, a pathway that can bypass lysosomal degradation and enable transcytosis (Ortega-Liebana and Sanchez-Martin, 2025). Positively charged QDs were internalized via non-Fc receptor-dependent pathways (e.g., macropinocytosis or caveolin-mediated endocytosis) (Liu Y.-Y. et al., 2025). Understanding these uptake pathways is critical because they dictate intracellular trafficking kinetics and the final destination of the drug cargo, thus directly influencing the therapeutic efficacy. Nonetheless, the clinical translation of cadmium-based QDs systems is constrained by persistent concerns regarding the long-term toxicity, primarily due to potential cadmium ion leakage and the associated heavy metal exposure.

A promising alternative lies in CQDs, which offer comparable optical properties with markedly improved biocompatibility, low/none toxicity, and environmental friendliness. The manner in which a therapeutic drug is loaded onto CQDs, whether via physical adsorption, electrostatic binding, or covalent conjugation, critically determines the release kinetics, stability, and overall therapeutic efficacy (Noorizadeh and Letters, 2025; Krishnaswamy and Pandian, 2022). Physical adsorption strategy refers to the method through which molecules are physically adsorbed on carriers via inter-molecule interactions, such as ionic interactions, H-bonds, van der Waals forces, hydrophobic interactions, and dipole–dipole interactions, which is due to the saturation of the counter-ions of the carriers or the fast ion exchange that cause the initial rapid drug release (Li Q. et al., 2020). Electrostatic binding, while reversible, is sensitive to ionic strength and pH changes, leading to premature drug loss (Dua et al., 2023). In contrast, covalent conjugation forms stable amide or thioether bonds (e.g., via polyacrylicacid/polyethylene glycol or thiol-maleimide chemistry), which minimize premature release and provide controlled release profiles (Pourmadadi et al., 2024; Bao et al., 2019). To further enhance specificity, stimuli-responsive linkers, such as pH-sensitive hydrazone bonds or reducible disulfide bonds, can be introduced, allowing drug release only upon exposure to the acidic tumor microenvironment (Phillips et al., 2014; Pillarisetti et al., 2017). Recent research has focused on designing intelligent CQD-based nanocarriers with stimuli-responsive behaviors. Feng et al. (2016a), Feng et al. (2016b) developed a pH-responsive, charge-convertible CQDs nanocarrier for the delivery of cisplatin (IV), and the prepared CDs-based drug nanocarrier enhanced in vivo cancer therapeutic efficiency. As shown in Figure 7, the drug is covalently conjugated via pH-sensitive linkages, ensuring minimal premature release during circulation and rapid release upon endosomal acidification. This system, termed CQDs-Pt(IV)@PEG-(PAH/DMMA), exploits the pH difference between the tumor extracellular microenvironment to trigger drug release. Its smart design in featuring a PEG functionalized poly(allylamine) shell with hydrolyzable dimethyl maleic acid (DMMA) groups prolongs drug circulation time, enhances tumor accumulation through the enhanced permeability and retention (EPR) effect, facilitates endosomal escape, and eventually enhances the therapeutic effect in vivo. The high surface-area-to-volume ratio of CQDs, high loading, combined with amphiphilic polymer coatings or hollow structures, further improves the drug-loading capacity and stability. Furthermore, CQDs were modified for surface functionalization with targeting ligands, which can direct to specific tissues. A prominent example is the use of transferrin conjugation to mediate the blood–brain barrier (BBB) penetration. It was reported that transferrin, a ligand, could bind to the transferrin receptors and ferry transferrin-conjugated carriers to the central nervous system (CNS) (Wiley et al., 2013; Roberts et al., 1993). In this study, transferrin, dye-labeled transferrin, or fluorescein were first covalently conjugated to CQDs and tested for their ability to cross the BBB. CQDs or the conjugates, transferrin-CQDs, and dye-transferrin-CQDs were injected intravascularly to the heart of the zebrafish to examine the possibility of transporting CQDs into the CNS. Results from the confocal fluorescence images’ fluorescence intensity were tested in the dye–transferrin–CQDs group but not in both the control and CQDs group. The fluorescence intensity was strong enough, and transferrin-conjugated CQDs could successfully “light up” the central nervous system (Li et al., 2016a). Li et al. (2016b) further designed a targeted delivery system (C-Dots-Trans-Dox) by conjugating doxorubicin to transferrin-coated CQDs. Four pediatric cell lines with overexpressed transferrin receptors and SJGBM2 cells were chosen for treatment with 500 nM conjugate or Dox to examine the cellular uptake. Results showed that approximately 5-fold more Dox was observed in the nuclei of the C-Dots-Trans-Dox group than in the group receiving Dox alone, indicating that Dox was delivered to these brain tumor cells effectively by the transferrin–CQDs-mediated delivery system. For the efficacy of C-Dots-Trans-Dox, in vitro experiments showed that C-Dots-Trans-Dox at 10 nM was significantly more cytotoxic than Dox alone, reducing viability by 14%–45%, across multiple pediatric brain tumor cell lines. Therefore, more research regarding the QDs–Dox micelle complex needs to be conducted to determine its efficacy and safety.

Concerns about the biocompatibility and long-term safety of QDs are prominent, particularly regarding those containing heavy metals such as cadmium. Research is essential to develop safer alternatives that are free from cadmium and other toxic materials, ensuring that QDs can be safely used in clinical settings (Mandal et al., 2013). To date, only 10 clinical trials for QDs have been registered, with one being completed and three being subsequently withdrawn (Table 3) (Pareek et al., 2025). Despite promising preclinical advances, several multifaceted challenges hinder the clinical adoption of QD-based drug delivery systems. First, the potential toxicity of QDs might compromise the patient’s health over time. The risks of ion leaching, oxidative stress, and cumulative organ toxicity necessitate rigorous in vivo chronic toxicity studies (Lin and Chen, 2023; Liu et al., 2022). Even for CQDs, some index such as detailed pharmacokinetics, biodistribution, immunogenicity, and potential off-target effects should be evaluated in relevant animal models (Ali et al., 2025; Kumar et al., 2025). Quantitative pharmacokinetic parameters including circulation half-life, volume of distribution, clearance rate, area under the curve, and mean residence time are essential for evaluating the clinical potential of QDs, but they remain largely unreported for most QDs formulations (Ishaque et al., 2025). For nanomaterials delivered intravenously, surface chemistry critically determines these parameters: half-life of the PEGylated nanoparticles increased from 1 to 241 min, whereas non-PEGylated or positively charged QDs are cleared within minutes (Mishra et al., 2025; Jednorski, 2023). For efficient renal clearance, QDs require an optimal diameter below 5.5 nm; while larger ones (>10 nm) predominantly accumulate in the liver and spleen, with over 80% of the injected dose often localized in these organs within 24 h (Ishaque et al., 2025). Such quantitative biodistribution data are critical for predicting the therapeutic efficacy and long-term safety. Second, QDs can undergo degradation, compromising their performance as drug-delivery vehicles (Hardman, 2005). Surface modification and encapsulation techniques are being explored to enhance these parameters. Heterogeneous expression of the target receptors in patient populations, non-specific uptake by the reticuloendothelial system, and physiological barriers can severely limit the delivery efficacy (Hu et al., 2024). Upon contact with biological fluids (blood, intestinal, or respiratory tract fluid), QDs that are rapidly covered with biomolecules, mainly protein, form a so-called protein corona, which can alter the physicochemical properties of QDs, thereby increasing the hydrodynamic diameter (Monopoli et al., 2020; Bilardo et al., 2022). Such aggregation accelerates reticuloendothelial system clearance, shortens circulation half-life, and alters the volume of distribution, leading to enhanced liver/spleen accumulation and reduced tumor targeting (Khosla et al., 2025; Wang Y. et al., 2025). Third, the interaction of QDs with the immune system represents another critical hurdle. Upon systemic administration, QDs are opsonized by plasma proteins, which tag them for recognition and uptake by macrophages of the reticuloendothelial system (Peng et al., 2024). Surface-engineering strategies such as dense PEGylation, zwitterionic coatings, or CD47-derived “do not eat me” peptides can inhibit phagocytosis and prolong circulation (Singh et al., 2026). However, multiple dosing of PEGylated drugs did not completely abolish protein adsorption and may even trigger anti-PEG antibodies, leading to accelerated clearance (Fu et al., 2025). Rational design should aim for controlled immune interactions that balance circulation time with eventual clearance and safety. Fourth, scalable manufacturing processes for the widespread clinical use of QDs should be established. Emerging methods, such as PDMS-based devices and additive-manufactured microfluidics, offer scalable, cost-effective, and sustainable production of QDs with enhanced application potential (Fadlelmula et al., 2024; bin Nasser Sarbaland et al., 2025). Addressing these challenges, we require sustained interdisciplinary collaboration across materials science, pharmacology, toxicology, and regulatory science, paving the way for the clinical realization of QDs-based theranostics.

TABLE 3. Clinical trials involving quantum dots.

3.3 Single-virus tracking

Viral infection is a complex and dynamic process involving specific receptor binding, cellular entry (often via endocytosis or membrane fusion), intracellular transport, and eventual release of the viral genome at appropriate sites for replication (Marsh and Helenius, 2006). Understanding these mechanisms of viruses is an important way to prevent and treat viral diseases and develop antiviral drugs. SVT has emerged as a crucial technique to follow individual viruses, visualize their transport behaviors, dissect their dynamic interactions with the host cells, and reveal the detailed insights into infection dynamics (Wang et al., 2021; Li et al., 2018). A key advantage of SVT is its ability to monitor these processes directly and dynamically over extended periods. Compared with traditional organic dyes, the excellent brightness and superior photostability of QDs enable them to be tracked for extended periods of time with low laser intensity, making them particularly favorable for acquiring time-series images or z-stacks for 3D reconstructions (Dahan et al., 2003; Grütter et al., 2006). In 2008, QDs were first successfully utilized to specifically label the envelope of viruses without significant effect on the infectivity of the virus, thereby allowing the uptake mechanism to be analyzed in detail (Joo et al., 2008). This was considered a watershed moment for SVT, with increasing technologies and methods being used for real-time tracking of viruses.

QDs have high brightness, which enables labeling a virus with sufficient brightness using very few QDs or even just one QD. Moreover, QDs hold the potential to facilitate the conjugation of a virus with a variety of labeling strategies because of the abundant surface ligands. As shown in Figure 8, many strategies, such as biotin–streptavidin interaction, direct chemical reaction, and peptide tag-mediated labeling have been developed to label virus with fluorescence by QDs-based single-virus tracking (QSVT). For instance, Hong et al. (2015) devised a convenient hydrazine-aldehyde-based strategy to label the influenza A virus (IAV) by conjugating 4-formylbenzoate-modified QDs to 6-hydrazinonicotinate acetone hydrazone-modified viral surface, achieving high selectivity, stable conjugation, and a labeling efficiency of 92%–93% without compromising infectivity. Building on surface labeling, the coupling of QDs to the viral ribonucleoprotein (vRNP) complexes inside IAV was developed, where vRNP complexes were central to every aspect of the IAV life cycle. In this study, as shown in Figure 9, green QDs-labeled nucleoprotein antibodies were transferred into H1N1 to achieve labeling of vRNP complexes with QDs, resulting in a two-color QD-labeled IAV, which could maintain 93% infectivity (Hong et al., 2018). Recently, the team of Dai-wen Pang (Du et al., 2024; Fu et al., 2024) further used QSTV to comprehensively visualize the role of the actin cytoskeleton in IAV entry and quantitatively dissected the triple roles of dynactin in dynein-driven transport. The results showed three different pathways for IAV to reach dynein along microtubules (MTs), namely, the virus (1) was driven by myosin VI to move along actin filaments to reach MTs (AF); (2) was propelled by actin tails assembled by an Arp2/3-dependent mechanism to reach MTs; and (3) directly reached MTs without experiencing actin-related movement (Du et al., 2024). After the entry of virus, virus-carrying vesicles bound to dynein and dynactin before being delivered to MTs. The attachment of dynein to the vesicles was dependent on dynactin and its subunits p150 Glued and Arp1 (Fu et al., 2024). Wen et al. (2014) proved a host cell-assisted strategy to label the nucleocapsid of real enveloped viruses with QDs. Briefly, VP39-AP was coupled with biotin and biotinylated nucleocapsid during the virus replication process, the virus was assembled, and it was finally budded to form the recombinant baculovirus (RBV). SA-QDs were subsequently used to site-specifically label the nucleocapsid of RBV based on the close interaction between biotin and streptavidin, monitored by tracking single-QDs-RBV in real-time. Zhang et al. (2020) reported a general method to mildly and readily label enveloped viruses, where lipid–biotin conjugates were used to recognize and mark viral lipid membranes and SA-QDs were used to light enveloped viruses, such as JEV, porcine reproductive and respiratory syndrome virus, and influenza A virus. Enveloped viruses were labeled in 2 h with specificity and efficiency up to 99% and 98%, respectively, and key protein-mutated in JEV did not affect the virus intracellular trafficking. These findings indicated that the streptavidin system was developed as a method for site-specifically labeling viral particles with QDs for single-virus tracking and provided important insights into the cell’s endocytic transport mechanism.

Packaging QDs into the internal components of viruses, such as DNA or RNA, and protein complexes is optimal for monitoring late infection events, enabling enhanced understanding of viral infection mechanisms. However, these approaches lack targeting and specificity. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system utilizes CRISPR RNA to accurately direct foreign nucleic acid recognition and targeting, which provides a potential strategy to label internal viral components with QDs’ specificity, reliability, and commonality (Yang et al., 2020a; Huang et al., 2023; Thuma et al., 2023). Yang et al. (2020a) firstly utilized a CRISPR imaging system to label PRV nucleic acids with QDs. As shown in Figure 10, biotinylated dCas9 protein and a gRNA against the US2 gene of PRV were designed and prepared in vitro. Then, dCas9-Bio/gRNAUS2 complexes and SA-QDs were transfected into HEK293T cells to form PRV-QD, followed by infection with PRV. Vero and HeLa cells were used to monitor the processes of PRV-QDs in real-time, including PRV-QDs adsorption, cytoplasmic transport along microtubules, and nuclear entry, indicating that this approach can be used for long-term SVT (Yang et al., 2020a; Yang et al., 2020b). Ma et al. (2024) utilized the CRISPR/Cas13 system and a bio-orthogonal metabolic method to label JEV genome and envelopes with different-colored QDs in situ. The combination of this procedure with STV aided in visualizing JEV uncoating and genome release in real-time near the endoplasmic reticulum of live cells. Therefore, QDSV is an exquisitely selective and powerful approach to study how viruses internalize in host cells dynamically and release their genome for viral replication and assembly, ensuring the completion of viral life cycles.

4 Biosafety of quantum dots

QDs can participate in cell imaging, drug-targeted therapy, and virus tracking, which can greatly improve the current medical status and provide targeted medical diagnosis and treatment. It is likely to become a core nanomaterial in the research of biomedical materials and has gained wide attention from researchers around the world. However, the toxicity of QDs, such as immunotoxicity, hepatotoxicity, nephrotoxicity, and respiratory toxicity, among others, still is the main factor that limits their applications. The toxicity mechanisms of QDs can be summed up as the following three points: (1) the core of QDs interact with the medium to degrade and release heavy metal ions such as Cd2+; (2) increased intracellular oxidative stress by producing ROS and inducing apoptosis; (3) non-specific adsorption to the surface of the cell, binding with the protein, destroying the structure, and affecting the normal function of the cell. The toxicity of QDs is a complex process, including the interaction with the body, cells, and molecules (Campalani and Monbaliu, 2025). The toxicity of QDs depend on multiple physicochemical factors, including the chemical composition, particle size, different surface modifications, exposure route, time and administration of QDs, and even the environmental condition (Wang and Tang, 2018; Lin and Chen, 2023). Therefore, scientists are making great efforts to solve this problem in order to use them safely for subsequent bioconjugation with proteins, peptides, or other chemical moieties.

4.1 Toxicity mechanisms

Before exploring the factors influencing the QDs’ toxicity in detail, it is essential to understand the primary mechanisms underlying their cytotoxic effects, which generally follow three interconnected pathways, namely, degradation-mediated release of toxic ions, generation of reactive oxygen species (ROS), and direct interaction with cellular membranes and intracellular components. First, the core of conventional QDs can interact with the medium to degrade and release heavy metal ions such as Cd2+ and Ag+ (Fan et al., 2023). Traditional QDs, including CdSe, CdTe, or CdS, contain cadmium—a heavy metal known for its potential cytotoxicity, genotoxicity, and carcinogenicity. Under physiological conditions, multiple factors including pH, ionic strength, and enzymatic activity can destabilize QDs, including surface degradation or corrosion of the inorganic core, leading to the leaching of Cd²⁺ ions (Sabarees, 2026; Katendra et al., 2026). Once released Cd²⁺ ions disrupt mitochondrial function by binding to sulfhydryl groups in key mitochondrial proteins, leading to oxidative stress and functional impairment. The rate and extent of ion release are highly dependent on environmental conditions: acidic environments (e.g., endosomes and lysosomes, pH 5.0) significantly accelerate Cd²⁺ release compared to neutral environments with pH (6.5) and pH (9.0), while high ionic strength promotes QD aggregation and alters the surface reactivity, thereby affecting degradation kinetics (Paydary, 2018; Mohammadpour and Molaabasi, 2023). Physiological concentrations of proteins and enzymes can bind to QD surfaces, either stabilizing them through corona formation or accelerating degradation via ligand displacement or enzymatic attack. Second, semiconductor QDs, particularly those containing heavy metals, can undergo photo-induced electron transfer upon exposure to light, leading to the production of ROS, such as superoxide, hydroxyl radicals, and singlet oxygen (Chen P. et al., 2026). This process occurs when photoexcited electrons or holes migrate to the QDs’ surface and react with molecular oxygen or water, respectively, and the oxidative stress thus caused can damage cellular components such as lipids, proteins, and DNA, ultimately triggering apoptosis or necrosis (Jomova et al., 2023; Zhao et al., 2023). Several factors influence ROS generation, including the core composition (e.g., CdTe produces more ROS than CdSe or CdS due to its narrower bandgap) and surface coating, (e.g., ZnS shells reduce ROS by confining excitons) (Chen Y. et al., 2026; Wang Z. et al., 2026). Importantly, ROS-mediated toxicity can occur even in the absence of heavy metal release, as demonstrated by carbon-based QDs (Chen P. et al., 2026; Famutimi et al., 2025). Third, another potential cause of QD toxicity is their direct interaction with cellular components. Some studies have indicated that QDs aggregate on the cell surface by binding to cell membrane receptors and are subsequently internalized via receptor-mediated or clathrin-mediated endocytosis (Zhang et al., 2011; Saulite et al., 2017). This interaction can lead to cell membrane damage, structural alterations in membrane proteins, and disruptions in the transport of substances across the membrane. Once internalized, QDs may accumulate in endolysosomal compartments, where the acidic pH can further promote degradation and ion release, creating a positive feedback loop that exacerbates toxicity (Wang Q. et al., 2026; Guha and Ghosh, 2022). Moreover, surface charge critically influences these membrane interactions: positively charged QDs tend to disrupt membrane integrity more severely than neutral or negatively charged ones, leading to increased cytotoxicity and hemolysis (Jiang et al., 2024; Niveria et al., 2023). In summary, the toxicity of QDs arises from a complex interplay of ion leaching, ROS generation, and direct membrane interaction mechanisms that are not mutually exclusive but often operate synergistically depending on the physicochemical properties of the QDs and their environmental context. A mechanistic understanding of these pathways is essential for designing safer QDs and accurately predicting their in vivo biosafety.

4.2 Factors that influence toxicity of quantum dots

4.2.1 Chemical composition

The fundamental factor of QDs’ toxicity is their composition. By far, the most commonly used quantum dots are Cd-based QDs, such as CdSe (cadmium selenide). Cd²⁺ has been identified as a carcinogen by the International Cancer Research Center and is recognized as a toxic heavy metal, which accumulates among the mesh endometrial system, such as the liver, spleen, lung, and kidney, and exhibits various immune responses and tissue damage. Winiarska-Mieczan and Kwiecień (2016) reported that more than 60% of the Cd²⁺ administered to rats accumulates in the liver. The relationship between the distribution percentage of Cd in the studied organs can be presented as liver > kidneys > brain > lungs > heart > spleen, and the liver is a major target of Cd injury. Latest research has indicated that the main mechanism of liver damage caused by Cd²⁺ in vitro is the binding of sulfhydryl groups in key mitochondrial proteins, leading to functional inactivation of these proteins, leading to oxidative stress, DNA impairment, mitochondrial dysfunction, autophagy induction, inflammatory cell infiltration, and fibrosis (Zhang Y. et al., 2023; Sun et al., 2023; Liu et al., 2023). Simultaneously, a layer of semiconductor shell was usually coated on the surface of QDs to reduce the leakage of Cd²⁺ and the formation of free ions, thus reducing the toxicity. To address this problem, some non-Cd QDs, such as InP/ZnS (Brunetti et al., 2013), ZnSe/ZnS (Nie et al., 2022), CQDs (Chung et al., 2021), and CuInS₂/ZnS (Lim et al., 2023), have recently been developed. Of course, surface modifications, such as different surface functional groups (e.g., COOH, NH₂, or OH) or surface charge on QDs, affect their biological behavior and toxic effects (Zheng et al., 2021).

4.2.2 Particle size

The distribution and metabolism of QDs varied with the type, dose, particle size, and surface coating. Previously, Huang et al. (2019) found that there was a size-dependent recognition in QDs, where the smaller QDs tended to accumulate in human breast carcinoma cells and caused significant toxicity. Consistently, it has been confirmed that smaller QDs have stronger toxic effects on the immune system as they are more likely to pass through immune cells (Bi et al., 2023). However, unlike in vitro cultured cells, researchers pointed out that the biodegradation and excretion of QDs in organs and tissues was related to the clearance time and the size of QDs. Su et al. (2011) pointed out that biodistribution of QDs was size-dependent. The QDs initially accumulated in the liver, QDs with larger sizes accumulated more quickly in the spleen, and the smaller QDs were absorbed by the kidney more easily. As is known, the glomerular filtration barrier has a “size cut-off” criterion, where the smaller QDs are rapidly excreted through the kidneys. Meanwhile, the QDs larger 6 nm–8 nm accumulate in the body and take a long time to clear (Du et al., 2017). Ge et al. (2017) systematically studied the biodistribution and clearance of the ultra-small Ag₂Se QDs (<3 nm) in mice. The results showed that Ag₂Se QDs could be cleared out at 168 h even at the high injection dosage of 150 mg/kg, and the blood circulation half-life was 0.25 h. Moreover, there are no abnormal pathological changes in the heart, liver, spleen, lung, kidney, and brain of the mice.

4.2.3 Different surface modifications

Surface function group modification of QDs is an important factor for their biological effects. Since the majority of QDs are composed of toxic substances, shells or macromolecules are often coated on the surface in order to reduce the biotoxicity. In addition, researchers hope that the QDs could be eliminated from the body over time, rather than being broken down or accumulated in the organs for a long time. Zheng et al. (2021) compared the in vivo toxicity of CdSe/ZnS QDs with -COOH and -NH₂ modification using zebrafish embryos. They found that the two QDs mainly attached to the embryo membrane and the eyes, yolk, and pericardium of zebrafish larva and led to a decrease in embryo movement, survival rate, and hatching rate and an increase in cardiac malformation and dysfunction. Moreover, the carboxyl QDs are more toxic, possibly due to the higher affinity and greater release, to embryos and larvae. Li L. et al. (2020) systematically evaluated the effects of different surface modifications on the biodistribution and toxicity of InP QDs. In this research study, three kinds of InP/ZnS QDs with different surface modifications, hQDs (QDs-OH), aQDs (QDs-NH₂), and cQDs (QDs-COOH), were analyzed for the biodistribution and in vivo toxicity of InP/ZnS QDs. Biodistribution of the three QDs was determined through cryosection fluorescence microscopy and ICP-MS analysis. These types of InP/ZnS QDs were rapidly distributed in the major organs of mice, mainly in the liver and spleen, and lasted for 28 days, and the results of H&E staining showed that no obvious histopathological abnormalities were observed in the main organs. No abnormal behavior, weight change, or organ index were observed during the whole observation period. These results provided a better understanding of the effects of the surface chemistry on the toxicity of QDs at the animal level and also facilitate the synthesis of safer QDs in biomedical applications.

5 Conclusions and perspective

5.1 Conclusion

This review article is based on the prospective and application of quantum dots in biomedical. A brief summary about the types of quantum dots, optical properties, as well as synthesis and characterization methods. Subsequently, applications of quantum dots in the biomedical field, such as cell imaging, drug delivery were discussed and with examples were covered. A particular emphasis focal on roles/applications for QDs-based single-virus tracking, which offered a powerful tool to monitor the dynamic behaviors, infection mechanisms of viruses. Finally, the toxicity mechanisms of quantum dots are discussed based on their chemical composition, particle size, and surface modifications.

Quantum dots are a new type of inorganic nanocarriers composed of heavy metal or inorganic core material coated with shell. Based on their core composition, QDs can be classified into 12 main categories: I B-VI A, I B-VII A, II B-VI A, III A-V A, IV A-VI A, IV A, V A, I B-III A-VI A, P dots, TMDCs, MXene, and Perovskite. The name quantum dots itself indicates its quantum confinement, optical properties and high surface-to-volume ratio. QDs can be synthesized through various methods, including colloidal synthesis, electrochemical synthesis, bio-templated synthesis, and biosynthesis. Each method is associated with certain advantages and disadvantages. Characterization of QDs can be done by spectro-scopic, microscopic, dynamic light scattering, electrical techniques and so on. Functionalization or surface modification of QDs made them to meet specific biomedical applications. Recent research about application of QDs in biomedical summarized here, there are still several major roadblocks. Here, we particularly discussed the QSTV technology. The tacking of the virus infection process not only needs biological labels with excellent performance but also requires the establishment of the efficient, specific, fixed-point, and quantitative in situ labeling methods according to the structure of the virus and the characteristics of the dynamic infection process, which is crucial to acquiring more accurate information by QSVT. So far, monitoring virus assembly and egress with QSVT remains a challenge. Most of the current SVT studies are carried out at the cellular level, which cannot provide the most authentic and reliable information for understanding the viral infection mechanisms. Thus, it is urgent to develop new labeling methods and imaging methodologies to probe the viral journey in live tissues or in vivo. With decades of development of QSVT in virological research, new strategies for virus labeling and cutting-edge imaging instrumentation and the establishment of suitable data analysis methods will be development to uncover unrevealed mechanisms of virus infection.

5.2 Future prospect

As toxicity concerned in biological field is the major issue with quantum dots; in future attempts would be made to minimize this toxicity with improved pharmacological response. Development of non-toxic QDs or functionalizing QDs to target specific organs or tissues may lower systemic toxicity. Information about the intracellular delivery of quantum dots is yet to be investigated because its bio distribution, metabolism, excretion, under the disease mode is not clear. Quantum dots thus have great future in medical field and is still a long process.

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Footnotes

Edited by: Kumud Malika Tripathi, Indian Institute of Petroleum and Energy (IIPE), India

Reviewed by: Jaya Dwivedi, Banasthali University, India

Himanshu Asati, Indian Institute of Technology Jodhpur, India