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After entering host cells by endocytosis, influenza A virus (IAV) is transported along microfilaments and then transported by dynein along microtubules (MTs) to the perinuclear region for genome release. Understanding the mechanisms of dynein-driven transport is significant for a comprehensive understanding of IAV infection. In this work, the roles of dynactin in dynein-driven transport of IAV were quantitatively dissected in situ using quantum dot-based single-virus tracking. It was revealed that dynactin was essential for dynein to transport IAV toward the nucleus. After virus entry, 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. Once viruses reached MTs, dynactin-assisted dynein initiates retrograde transport of IAV. Importantly, the retrograde transport of viruses could be initiated at both plus ends (32%) and other regions on MTs (68%). Subsequently, dynactin accompanied and assisted dynein to persistently transport the virus along MTs in the retrograde direction. This study revealed the dynactin-dependent dynein-driven transport process of IAV, enhancing our understanding of IAV infection and providing important insights into the cell's endocytic transport mechanism.

Developing the second near-infrared (NIR-II) photoacoustic (PA) agent is of great interest in bioimaging. Ag 2 Se quantum dots (QDs) are one kind of potential probe for applications in NIR-II photoacoustic imaging (PAI). However, the surfaces with excess anions of Ag 2 Se QDs, which increase the probability of nonradiative transitions of excitons benefiting PA imaging, are not conducive to binding electron donor ligands for potential biolabeling and imaging. In this study, Staphylococcus aureus (S. aureus) cells are driven for the biosynthesis of Ag 2 Se QDs with catalase (CAT). Biosynthesized Ag 2 Se (bio-Ag 2 Se-CAT) QDs are produced in Se-enriched environment of S. aureus and have a high Se-rich surface. The photothermal conversion efficiency of bio-Ag 2 Se-CAT QDs at 808 and 1064 nm is calculated as 75.3% and 51.7%, respectively. Additionally, the PA signal responsiveness of bio-Ag 2 Se-CAT QDs is ≈10 times that of the commercial PA contrast agent indocyanine green. In particular, the bacterial CAT is naturally attached to bio-Ag 2 Se-CAT QDs surface, which can effectively relieve tumor hypoxia. The bio-Ag 2 Se-CAT QDs can relieve heat-initiated oxidative stress while undergoing effective photothermal therapy (PTT). Such biosynthesis method of NIR-II bio-Ag 2 Se-CAT QDs opens a new avenue for developing multifunctional nanomaterials, showing great promise for PAI, hypoxia alleviation, and PTT., (© 2024 Wiley‐VCH GmbH.)

Dopamine (DA) is an important small-molecule neurotransmitter, which is closely related to the development of many neurological diseases and has received increasing attention in the diagnosis of neurological diseases. Currently, the assays of the detection of dopamine such as electrochemical and colorimetric methods have low sensitivity, poor selectivity and susceptibility to interference, which limit the accurate quantification of dopamine. Fluorescence anisotropy immunoassay is a traditional analytical method in which the quantification is based on the change in fluorescence anisotropy values observed when fluorescence molecules are bound to a certain volume and mass of the material. Since dopamine is a small molecule with small volume and mass, we took advantage of the good photostability of the second near-infrared window (NIR-II) quantum dots (QDs) and the low spontaneous interference of the substrate, and designed a biosensor dopamine fluorescence anisotropy probe streptavidin biosensor (DFAP-SAB) based on the NIR-II QDs combined with streptavidin signal amplification to achieve rapid and separation-free detection of dopamine in human serum. The detection signal has a good linearity between 50 nM and 3000 nM with a detection limit of 11.2 nM. The application of NIR-II QDs provides the possibility of biosensor applications for complex samples. The construction of the streptavidin signal amplification device provides a new idea for small molecule detection.

In this work, a novel silver ion (Ag + )-regulated ratiometric fluorescence method for the effective and sensitive determination of alkaline phosphatase (ALP) was established based on carbon dots (CDs) and o-phenylenediamine (OPD). OPD can be oxidized by Ag + to generate fluorescent 2, 3-diaminophenazine (DAP). Thus, based on inner-filter effect (IFE) or/and fluorescence resonance energy transfer (FRET) between CDs and DAP, the CDs-Ag + -OPD system can generate dual-emission at 454 nm and 570 nm respectively when excited at 360 nm. The introduction of ascorbic acid (AA) can react with Ag + to produce dehydroascorbic acid (DHAA), which inhibits the generation of DAP, resulting in the fluorescence decrease at 570 nm and fluorescence recovery of CDs at 454 nm. Meanwhile, DHAA can react with OPD to generate quoxaline (QX), which emits strong blue fluorescence at 440 nm, further inhibiting the IFE or/and FRET between CDs and DAP. An obvious ratiometric fluorescence response was observed with the increase of the concentration of AA introduced. Due to the fact that AA can be generated by the enzyme catalysis reaction between ALP and 2-phospho-l-ascorbic acid (AAP), the CDs-Ag + -OPD ratiometric system was applied to the determination of ALP successfully. The ratiometric fluorescence value of F 454 /F 570 increases with increasing ALP concentration, with a linear range of 0.2 to 40 U/L and detection limit of 0.1 U/L. In addition, the CDs-Ag + -OPD ratiometric system was successfully applied to the detection of ALP in human serum samples., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

The surface of nanocrystals plays a dominant role in many of their physical and chemical properties. However, controllability and tunability of nanocrystal surfaces remain unsolved. Herein, we report that the surface chemistry of nanocrystals, such as near-infrared Ag 2 Se quantum dots (QDs), is size-dependent and composition-tunable. The Ag 2 Se QDs tend to form a stable metal complex on the surface to minimize the surface energy, and therefore the surface chemistry can be varied with particle size. Meanwhile, changes in surface inorganic composition lead to reorganization of the surface ligands, and the surface chemistry also varies with composition. Therefore, the surface chemistry of Ag 2 Se QDs, responsible for the photoluminescence (PL) quantum yield and photostability, can be tuned by changing their size or composition. Accordingly, we demonstrate that the PL intensity of the Ag 2 Se QDs can be tuned reversely by adjusting the degree of surface Ag + enrichment via light irradiation or the addition of AgNO 3 . This work provides insight into the control of QD surface for desired PL properties., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2021 Science China Press. Published by Elsevier B.V. All rights reserved.)

A novel ratiometric fluorescence nanoprobe based on long-wavelength emission carbon dots (CDs) was designed for high sensitive and selective detection of Zn 2+ . The CDs were conveniently prepared by a one-step solvothermal treatment of formamide and glutathione (GSH). Under single excitation wavelength (420 nm), the obtained CDs exhibit three emission peaks at 470, 650, and 685 nm, respectively. For the long-wavelength emission region of the CDs, the fluorescence at 685 nm can be quenched with different levels upon the addition of most metal ions. However, the presence of Zn 2+ not only results in the fluorescence quenching at 685 nm effectively but also enhances at 650 nm remarkably, which may be due to the formation of CD-Zn 2+ chelate complex inducing the dispersion of CDs aggregates and changes in the group distribution on the surface of CDs. Taking the advantage of the unique fluorescence response induced by Zn 2+ , the prepared CDs were successfully employed as nanoprobe for self-ratiometric fluorescence determination of Zn 2+ with F 650 /F 685 as signal output. A good linear relationship in the concentration range 0.01 to 2 μM, and a detection limit as low as 5.1 nM has been obtained. The ratiometric nanoprobe was successfully applied to  Zn 2+ determination  in human serum samples., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Fluorescence anisotropy (FA) has been widely applied for detecting and monitoring special targets in life sciences. However, matrix autofluorescence restricted its further application in complex biological samples. Herein, we report a near-infrared-II (NIR-II) FA strategy for detecting adenosine triphosphate (ATP) in human serum samples and breast cancer cell lysate, which employed NIR-II fluorescent Ag 2 Se quantum dots (QDs) as tags to reduce matrix autofluorescence effect and applied graphene oxide (GO) to enhance fluorescence anisotropy signals. In the presence of ATP, the recognition between NIR-II Ag 2 Se QDs labeled aptamer (QD-pDNA) and ATP led to the release of QD-pDNA from GO, resulting in the obvious decrease of FA values. ATP could be quantitatively detected in concentrations ranged from 3 nM to 2500 nM, with a detection limit down to 1.01 nM. This study showed that the developed NIR-II FA strategy could be applied for detecting targets in complex biological samples and had great potential for monitoring interactions between biomolecules in biomedical research., (Copyright © 2020 Elsevier B.V. All rights reserved.)

We herein used Ag 2 Se quantum dots (QDs) as a target-modulated sensitizer for upconversion nanoparticles (UCNPs) and the target thrombin as the sensitizing switch to construct a biosensor, circumventing the limited luminescence resonance energy transfer (LRET) efficiency of UCNPs, with enhanced signal-to-background ratio (SBR) and assay sensitivity.

The accurate and rapid monitoring of the expression levels of enterovirus 71 (EV71)-related microRNAs (miRNAs) can contribute to diagnosis of hand, foot, and mouth disease (HFMD) at the early stage. However, there is currently a lack of convenient methods for simultaneous monitoring of multiplex miRNAs in one step. Herein a one-step method for the simultaneous monitoring of multiple EV71 infection-related miRNAs is developed based on core-satellite structure assembled with magnetic nanobeads and quantum dots (MNs-ssDNA-QDs). In the presence of target miRNAs, duplex-specific nuclease (DSN)-assisted target recycling can be triggered, resulting in the release of QDs and recycling of target miRNAs. Then the simultaneous quantification can be easily realized by recording the corresponding amplified fluorescence signal of QDs in the suspension. With this method, simultaneous detection of hsa-miRNA-296-5p and hsa-miRNA-16-5p, potential biomarkers of EV71 infection, can be easily achieved with femtomolar sensitivity and single-base mismatch specificity. Moreover, the method is successfully used for monitoring of the expression level of miRNAs in EV71-infected cells at different time points, demonstrating the potential for diagnostic applications. With the merits of one-step operation and single-nucleotide mismatch discrimination, this work opens a new avenue for multiplex miRNAs detection. As different nucleotide sequences and multicolor QDs can be employed, this work is expected to offer great potential for the development of high throughput diagnosis.

Non-toxic and long-term fluorescent probes for tumor imaging are in urgent need for non-invasively obtaining information about tumor genesis and metastasis in vivo. Here, we present a biocompatible near-infrared fluorescent probe for in vivo long-term imaging of tumor by modifying glucose (Glc), which experiences high uptake in cancer cells, on the surface of near-infrared Ag 2 Se quantum dots (NIR Ag 2 Se QDs). The fluorescence of glucose-functionalized Ag 2 Se QDs (Glc-Ag 2 Se QDs) from the targeted tumor can be observed in vivo for at least 7 days. In addition, this probe could be excreted through kidneys and the renal excretion ability is favorable for in vivo imaging applications. Moreover, Glc-Ag 2 Se QDs could be used for tumor targeted imaging of not only human breast cancer cells (MCF-7), but also SW1990 pancreatic cancer cells since glucose is highly taken up in almost all kinds of tumors. Glc-Ag 2 Se QDs could be a promising general tool for in vivo long-term observation of tumor evolution.

Nanosized oncolytic viral light particles (L-particles), separated from progeny virions, are composed of envelopes and several tegument proteins of viruses, free of nucleocapsids. The noninfectious L-particles experience the same internalization process as mature oncolytic virions, which exhibits great potential to act as targeted therapeutic platforms. However, the clinical applications of L-particle-based theranostic platforms are rare due to the lack of effective methods to transform L-particles into nanovectors. Herein, a convenient and mild strategy has been developed to transform L-particles into near-infrared (NIR) fluorescence Ag 2 Se quantum dot (QD)-labeled active tumor-targeting nanovectors for real-time in situ imaging and drug delivery. Utilizing the electroporation technique, L-particles can be labeled with ultrasmall water-dispersible NIR fluorescence Ag 2 Se QDs with a labeling efficiency of ca . 85% and loaded with antitumor drug with a loading efficiency of ca . 87%. Meanwhile, by harnessing the infection mechanism of viruses, viral L-particles are able to recognize and enter tumor cells without further modification. In sum, a trackable and actively tumor-targeted theranostics nanovector can be obtained efficiently and simultaneously. Such multifunctional nanovectors transformed from viral L-particles have exhibited excellent properties of active tumor-targeting, in vivo tumor imaging, and antitumor efficacy, which opens a new window for the development of natural therapeutic nanoplatforms.

The advantages of fluorescence bioimaging in the second near-infrared (NIR II, 1000-1700 nm) window are well known; however, current NIR II fluorescent probes for in vivo tumor imaging still have many shortcomings, such as low fluorescence efficiency, unstable performance under in vivo environments, and inefficient enrichment at tumor sites. In this study, Ag 2 Te quantum dots (QDs) that emit light at a wavelength of 1300 nm are assembled with poly(lactic-co-glycolic acid) and further encapsulated within cancer cell membranes to overcome the shortcomings mentioned above. The as-prepared ≈100 nm biomimetic nanobioprobes exhibit ultrabright (≈60 times greater than that of free Ag 2 Te QDs) and highly stable (≈97% maintenance after laser radiation for 1 h) fluorescence in the NIR II window. By combining the active homotypic tumor targeting capability derived from the source cell membrane with the passive enhanced permeation and retention effect, improved accumulation at tumor sites ((31 ± 2)% injection dose per gram of tumor) and a high tumor-to-normal tissue ratio (13.3 ± 0.7) are achieved. In summary, a new biomimetic NIR II fluorescent nanobioprobe with ultrabright and stable fluorescence, homotypic targeting and good biocompatibility for enhanced in vivo tumor imaging is developed in this study., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The rapid and sensitive detection of pathogens is extremely crucial for timely clinical diagnosis and diseases control. Here, by employing cellular beacons with in situ synthesized QDs created from Staphylococcus aureus ( S. aureus), we efficiently fabricated an antibody (Ab) and acetylcholinesterase (AChE)-functionalized nanobioprobe, i.e., multifunctional cellular beacons (MCBs), avoiding complicated modification. Coupled with magnetic separation, a novel method for pathogen detection with the naked eye is established. With this method, enterovirus 71 (EV71) can be detected by the naked eye through the aggregation of gold nanoparticles that is triggered by the product of AChE catalyzed acetylthiocholine, with a detection limit of 0.5 ng/mL. Moreover, due to the MCBs have high luminance with perfect uniformity, the detection can also be realized by counting the number of MCBs, with a detection limit of 1 ng/mL. The method is validated with human throat swabs, resulting in a complete consistence with reverse transcription-polymerase chain reaction results. This study reports the first cellular beacons-based method for pathogen detection by the naked eye and broadens the applicability of cell self-synthesized nanoparticles-based immunoassays. Moreover, the MCBs-based method will provide a powerful tool for clinical detection.

Influenza A virus (IAV), a risk to public health, is enveloped and contains viral ribonucleoprotein (vRNP) complexes, where vRNP complexes are central to every aspect of the IAV life cycle. Labeling both the vRNP complexes and viral envelope with quantum dots (QDs) is conducive to achieving global long-term tracking of a single IAV infecting host cell, which has potential to provide valuable information for revealing mechanisms of IAV infection. However, even though some strategies for labeling of the viral envelope with QDs have been developed, there are few strategies for coupling of QDs to the vRNP complexes inside IAV so far. Herein, we devised a convenient electroporation-based strategy, coupled with antibody binding, to transfer green QDs-labeled nucleoprotein antibodies (GQDs-NPAb) into H1N1 and achieved the labeling of vRNP complexes with QDs [H1N1(GQDs)]. Under the optimal condition of 20 nM GQDs-NPAb and a single pulse with 20 ms duration and 750 V/cm pulse intensity, the actual efficiency of labeling is ca. 34% and H1N1(GQDs) can retain 93% infectivity. Then, dual labeling of H1N1 was realized by labeling the envelope of H1N1(GQDs) with red QDs (RQDs) via a mild and efficient hydrazine-aldehyde-based strategy. At the optimal RQDs concentration of 5 nM, the actual efficiency of dual labeling can reach to 11% and the dual-labeled H1N1 can retain 93% infectivity. Because of the similar components and structure of different IAV subtypes, this dual-labeling strategy is applicable to other subtypes of IAV, e.g., H9N2.

With the aid of the single-virus tracking technique and the quantum dot-labeling strategy, the internalization of the pseudorabies virus (PrV) has been visualized in real time. The results demonstrate that macropinocytosis can be considered as a major pathway for PrV entering HeLa cells, which facilitates the development of therapeutics for virus-triggered diseases.

Near infrared (NIR) emitting semiconductor quantum dots can be excellent fluorescent nanoprobes, but the poor biodegradability and potential toxicity limits their application. The authors describe a fluorescent system composed of graphene quantum dots (GQDs) as NIR emitters, and novel MnO 2 nanoflowers as the fluorescence quenchers. The system is shown to be an activatable and biodegradable fluorescent nanoprobe for the "turn-on" detection of intracellular glutathione (GSH). The MnO 2 -GQDs nanoprobe is obtained by adsorbing GQDs onto the surface of MnO 2 nanoflowers through electrostatic interaction. This results in the quenching of the NIR fluorescence of the GQDs. In the presence of GSH, the MnO 2 -GQDs nanoprobe is degraded and releases Mn 2+ and free GQDs, respectively. This gives rise to increased fluorescence. The nanoprobe displays high sensitivity to GSH and with a 2.8 μM detection limit. It integrates the advantages of NIR fluorescence and biodegradability, selectivity, biocompatibility and membrane permeability. All this makes it a promising fluorescent nanoprobe for GSH and for cellular imaging of GSH as shown here for the case of MCF-7 cancer cells. Graphical abstract A biodegradable NIR fluorescence nanoprobe (MnO 2 -GQDs) for the "turn-on" detection of GSH in living cell was established, with the NIR GQD as the fluorescence reporter and the MnO 2 nanoflower as the fluorescence quencher.

Autophagy is closely related to virus-induced disease and a comprehensive understanding of the autophagy-associated infection process of virus will be significant for developing more effective antiviral strategies. However, many critical issues and the underlying mechanism of autophagy in virus entry still need further investigation. Here, this study unveils the involvement of autophagy in influenza A virus entry. The quantum-dot-based single-virus tracking technique assists in real-time, prolonged, and multicolor visualization of the transport process of individual viruses and provides unambiguous dissection of the autophagic trafficking of viruses. These results reveal that roughly one-fifth of viruses are ferried into cells for infection by autophagic machineries, while the remaining are not. A comprehensive overview of the endocytic- and autophagic-trafficking process indicates two distinct trafficking pathway of viruses, either dependent on Rab5-positive endosomes or autophagosomes, with striking similarities. Expressing dominant-negative mutant of Rab5 suggests that the autophagic trafficking of viruses is independent on Rab5. The present study provides dynamic, precise, and mechanistic insights into the involvement of autophagy in virus entry, which contributes to a better understanding of the relationship between autophagy and virus entry. The quantum-dot-based single-virus tracking is proven to hold promise for autophagy-related fundamental research., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Quantum dots (QDs) are semiconducting nanocrystals with unique optical properties. When coated with shell/capping, QDs are not deleterious to cells and organisms. However, when QDs are retained in the cellular environment for a certain period of time, their coatings may be degraded, yielding "naked" QDs. Although some studies have documented the acute effects of cadmium telluride (CdTe) QDs in various cell lines, however, to our knowledge, there are no published studies on the chronic effects of CdTe QDs in normal lung cells. In this study, we therefore sought to study the effects of CdTe QDs of various particle sizes on their cytotoxicity and carcinogenicity in normal human bronchial epithelial cells (BEAS-2B). A total of three particle sizes of CdTe QD with emission maximum at 520, 580, and 730 nm were employed (abbreviated as 520Q, 580Q, and 730Q, respectively). Our results indicated that acute exposure to 520Q (∼2.04 nm in diameter) and 580Q (∼3.24 nm in diameter) elicited dose-dependent cytotoxicity; while acute exposure to 730Q (∼5.40 nm in diameter) elicited negligible cytotoxicity in BEAS-2B cells. Notably, chronic exposure to CdTe QD of all three tested particle sizes induced BEAS-2B cell transformation as evidenced by enhanced cell migration and anchorage-independent growth on soft agar. Taken together, our findings suggest that CdTe QDs are potent human lung carcinogens., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Quantum dot (QD)-based single virus tracking has become a powerful tool for dissecting virus infection mechanism. However, only virus behaviors at the early stage of retrograde trafficking have been dynamically tracked so far. Monitoring of comprehensive virus retrograde transportation remains a challenge., Results: Based on the superior fluorescence properties of QDs and their labeling of virus internal component, the dynamic interactions between baculoviruses and all key transportation-related cellular structures, including vesicles, acidic endosomes, actins, nuclear pores and nuclei, were visualized at the single-virus level. Detailed scenarios and dynamic information were provided for these critical interaction processes., Conclusions: A comprehensive model of baculovirus retrograde trafficking involving virus endocytosis, fusion with acidic endosome, translocation to nuclear periphery, internalization into nucleus, and arriving at the destination in nucleus was proposed. Thus the whole retrograde transportation of baculovirus in live host cells was elucidated at the single-virus level for the first time.

Ascorbic acid (AA), as one of the most important vitamins, participates in various physiological reactions in the human body and is implicated with many diseases. Therefore, the development of effective methods for monitoring the AA level in living systems is of great significance. Up to date, various technologies have been developed for the detection of AA. However, few methods can realize the direct detection of endogenous AA in living cells. In this work, we for the first time reported that near-infrared (NIR) graphene quantum dots (GQD) possessed good two-photon fluorescence properties with a NIR emission at 660 nm upon exciting with 810 nm femtosecond pulses and a two-photon (TP) excitation action cross-section (δΦ) of 25.12 GM. They were then employed to construct a TP nanoprobe for detection and bioimaging of endogenous AA in living cells. In this nanosystem, NIR GQDs (NGs), which exhibited lower fluorescence background in living system to afford improved fluorescence imaging resolution, were acted as fluorescence reporters. Also CoOOH nanoflakes were chosen as fluorescence quenchers by forming on the surface of NGs. Once AA was introduced, CoOOH was reduced to Co 2+ , which resulted in a "turn-on" fluorescence signal of NGs. The proposed nanoprobe demonstrated high sensitivity toward AA, with the observed LOD of 270 nM. It also showed high selectivity to AA with excellent photostability. Moreover, the nanoprobe was successfully used for TP imaging of endogenous AA in living cells as well as deep tissue imaging.

Highly efficient labeling of viruses with quantum dots (QDs) is the prerequisite for the long-term tracking of virus invasion at the single virus level to reveal mechanisms of virus infection. As one of the structural components of viruses, viral envelope lipids are hard to be labeled with QDs due to the lack of efficient methods to modify viral envelope lipids. Moreover, it is still a challenge to maintain the intactness and infectivity of labeled viruses. Herein, a mild method has been developed to label viral envelope lipids with QDs by harnessing the biotinylated lipid-self-inserted cellular membrane. Biotinylated lipids can spontaneously insert in cellular membranes of host cells during culture and then be naturally assembled on progeny Pseudorabies virus (PrV) via propagation. The biotinylated PrV can be labeled with streptavidin-conjugated QDs, with a labeling efficiency of ∼90%. Such a strategy to label lipids with QDs can retain the intactness and infectivity of labeled viruses to the largest extent, facilitating the study of mechanisms of virus infection at the single virus level., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Due to excellent optical properties, quantum dots (QDs) have been widely applied to sensing, labeling, and imaging. For the fabrication of QD-based bioprobes, purification is usually the crucial step. Hydrophilic octylamine grafted polyacrylic acid modified QDs (OPA-QDs) were prepared, and purified by high-performance size exclusion chromatography (HPSEC) to remove excess OPA and aggregated QDs. The percentage of suspended agglomerates of OPA-QDs in the unpurified OPA-QDs increases from 4% to 31% through a year, but the purified OPA-QDs of the same batch possess excellent colloidal stability for at least one year. Subsequently, QD-based bioprobes were fabricated by the conjugation between QDs and streptavidin (SA) or antibody (IgG), generating QD-SA and QD-IgG, respectively, which were purified via HPSEC. Finally, the resulting QD-SA and QD-IgG were adopted to detect tumour markers on slices and showed specific positive signals without nonspecific adsorption, which was contrary to the unpurified QD-IgG. Thus, the HPSEC-coupled system proposed in the current work is potent and universal for the generation of purified and monodisperse QD-based bioprobes, which is promising in the nanobiodetection field., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Ultrabright carbon nanodots-hybridized silica nanospheres (CSNs) are synthesized through the Stöber process of silane functionalized C-dots. The fluorescence of carbon nanodots is converged intensely. A CSN is about 3800 times brighter than a single-carbon nanodot. Along with their high brightness and low cytotoxicity, CSNs also indicate their potential application in cellular labeling., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The study of cancer is of great significance to human survival and development, due to the fact that cancer has become one of the greatest threats to human health. In recent years, the rapid progress of nanoscience and nanotechnology has brought new and bright opportunities to this field. In particular, the applications of quantum dots (QDs) and magnetic nanoparticles (MNPs) have greatly promoted early diagnosis and effective therapy of cancer. In this review, we focus on fluorescent/magnetic micro/nano-spheres based on QDs and/or MNPs (we may call them "nanoparticle-sphere (NP-sphere) composites") from their preparation to their bio-application in cancer research. Firstly, we outline and compare the main four kinds of methods for fabricating NP-sphere composites, including their design principles, operation processes, and characteristics (merits and limitations). The NP-sphere composites successfully inherit the unique fluorescence or magnetic properties of QDs or MNPs. Moreover, compared with the nanoparticles (NPs) alone, the NP-sphere composites show superior properties, which are also discussed in this review. Then, we summarize their recent applications in cancer research from three aspects, that is: separation and enrichment of target tumor cells or biomarkers; cancer diagnosis mainly through medical imaging or tumor biomarker detection; and cancer therapy via targeted drug delivery systems. Finally, we provide some perspectives on the future challenges and development trends of the NP-sphere composites.

Labeling and imaging of live cells with quantum dots (QDs) has attracted great attention in the biomedical field over the past two decades. Maintenance of the fluorescence of QDs in a biological environment is crucial for performing long-term cell tracking to investigate the proliferation and functional evolution of cells. The cell-penetrating peptide transactivator of transcription (TAT) is a well-studied peptide to efficiently enhance the transmembrane delivery. Here, we used TAT peptide-conjugated QDs (TAT-QDs) as a model system to examine the fluorescence stability of QDs in live cells. By confocal microscopy, we found that TAT-QDs were internalized into cells by endocytosis, and transported into the cytoplasm via the mitochondria, Golgi apparatus, and lysosomes. More importantly, the fluorescence of TAT-QDs in live cells was decreased mainly by cell proliferation, and the low pH value in the lysosomes could also lower the fluorescence intensity of intracellular QDs. Quantitative analysis of the amount of QDs in the extracellular region and whole cells indicated that the exocytosis was not the primary cause of fluorescence decay of intracellular QDs. This work facilitates a better understanding of the fluorescence stability of QDs for cell imaging and long-term tracking in live cells. Also, it provides insights into the utility of TAT for transmembrane transportation, and the preparation and modification of QDs for cell imaging and tracking.

Single-virus tracking (SVT) technique, which uses microscopy to monitor the behaviors of viruses, is a vital tool to study the real-time and in situ infection dynamics and virus-related interactions in live cells. To make SVT a more versatile tool in biological research, the researchers have developed a quantum dot (QD)-based SVT technique, which can be utilized for long-term and highly sensitive tracking in live cells. In this review, we describe the development of a QD-based SVT technique and its biological applications. We first discuss the advantage of QDs as tags in the SVT field by comparing the conventional tags, and then focus on the implementation of QD-based SVT experiments, including the QD labeling strategy, instrumentation, and image analysis method. Next, we elaborate the recent advances of QD-based SVT in the biological field, and mainly emphasize the representative examples to show how to use this technique to acquire more meaningful biological information.

Cell-derived microvesicles (MVs) are natural carriers that can transport biological molecules between cells, which are expected to be promising delivery vehicles for therapeutic purposes. Strategies to label MVs are very important for investigation and application of MVs. Herein, ultrasmall Mn-magnetofunctionalized Ag2Se quantum dots (Ag2Se@Mn QDs) integrated with excellent near-infrared (NIR) fluorescence and magnetic resonance (MR) imaging capabilities have been developed for instant efficient labeling of MVs for their in vivo high-resolution dual-mode tracking. The Ag2Se@Mn QDs were fabricated by controlling the reaction of Mn(2+) with the Ag2Se nanocrystals having been pretreated in 80 °C NaOH solution, with an ultrasmall size of ca. 1.8 nm, water dispersibility, high NIR fluorescence quantum yield of 13.2%, and high longitudinal relaxivity of 12.87 mM(-1) s(-1) (almost four times that of the commercial contrast agent Gd-DTPA). The ultrasmall size of the Ag2Se@Mn QDs enables them to be directly and efficiently loaded into MVs by electroporation, instantly and reliably conferring both NIR fluorescence and MR traceability on MVs. Our method for labeling MVs of different origins is universal and free of unfavorable influence on intrinsic behaviors of MVs. The complementary imaging capabilities of the Ag2Se@Mn QDs have made the long-term noninvasive whole-body high-resolution dual-mode tracking of MVs in vivo realized, by which the dynamic biodistribution of MVs has been revealed in a real-time and in situ quantitative manner. This work not only opens a new window for labeling with QDs, but also facilitates greatly the investigation and application of MVs.

Real-time tracking of fluorophore-tagged viruses in living cells can help uncover virus infection mechanisms. Certainly, the indispensable prerequisite for virus-tracking is to label viruses with some bright and photostable beacons such as quantum dots (QDs) via an appropriate labeling strategy. Herein, we devise a convenient hydrazine-aldehyde based strategy to label viruses with QDs through the conjugation of 4-formylbenzoate (4FB) modified QDs to 6-hydrazinonicotinate acetone hydrazone (HyNic) modified viruses under mild conditions. On the basis of this strategy, viruses can be successfully labeled with QDs with high selectivity, stable conjugation, good reproducibility, high labeling efficiency of 92-93% and maximum retention of both fluorescence properties of QDs and infectivity of viruses, which is very meaningful to tracking and statistical analysis of virus infection processes. By further comparing with the most widely used labeling strategy based on the Biotin-SA system, this new strategy has advantages of both high labeling efficiency and good retention of virus infectivity, thus offering a promising alternative for virus-labeling. Moreover, due to the ubiquitous presence of exposed amino groups on the surface of various viruses, this selective, efficient, reproducible and biofriendly strategy should have good universality for labeling both enveloped and nonenveloped viruses.

Avian influenza A(H7N9) virus, which emerged in China in the spring of 2013, has infected hundreds of people and resulted in many deaths. Herein, a rapid and quantitative assay is proposed for the one-step detection of H7N9 virions. Immunomagnetic nanospheres (IMNs) and antibody-conjugated quantum dots (Ab-QDs) are simultaneously employed to capture and identify the target virus, leading to a high efficiency, good specificity, and strong anti-interference ability. Moreover, this reliable detection assay, which combines the efficient magnetic enrichment and the unique photophysical properties of QDs, can achieve a high sensitivity for a low detection limit. At the same time, this detection strategy shows great flexibility for employment in a variety of fluorescence detectors, including fluorescence spectrometry, microscope assays, and handheld UV lamp tests. Furthermore, our one-step detection strategy induces very little change in the integrity of the vulnerable virions, which enables additional genotyping testing following the fluorescence detection. The present study, thus, reports a rapid and quantitative approach for the detection of H7N9 virions based on simultaneous magnetic capture and QD labeling, thereby providing a higher probability for detection and therefore faster diagnosis of H7N9-infected patients., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The photoluminescence of carbon nanodots (C-dots) can be tuned by changing their surface chemistry or size because the photoluminescence is a function of the surface-state electronic transitions. Increasing the degree of surface oxidation leads to a narrowing of the energy gap of the surface; meanwhile, larger C-dots with an extensive π-electron system, which can couple with surface electronic states, can also lead to a narrowing of the energy gap of the surface states., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Cell-derived microparticles (MPs) have been recently recognized as critical intercellular information conveyors. However, further understanding of their biological behavior and potential application has been hampered by the limitations of current labeling techniques. Herein, a universal donor-cell-assisted membrane biotinylation strategy was proposed for labeling MPs by skillfully utilizing the natural membrane phospholipid exchange of their donor cells. This innovative strategy conveniently led to specific, efficient, reproducible, and biocompatible quantum dot (QD) labeling of MPs, thereby reliably conferring valuable traceability on MPs. By further loading with small interference RNA, QD-labeled MPs that had inherent cell-targeting and biomolecule-conveying ability were successfully employed for combined bioimaging and tumor-targeted therapy. This study provides the first reliable and biofriendly strategy for transforming biogenic MPs into functionalized nanovectors., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Manipulating biochemical reactions in living cells to synthesize nanomaterials is an attractive strategy to realize their synthesis that cannot take place in nature. Yeast cells have been skillfully utilized to produce desired nanoparticles through spatiotemporal coupling of intracellular nonrelated biochemical reaction pathways for formation of fluorescent CdSe quantum dots. Here, we have successfully transformed Staphylococcus aureus cells into cellular beacons (fluorescing cells), all of which are highly fluorescent and photostable with perfect uniformity. Importantly, on the basis of such cells, we efficiently fabricated fluorescent nanobioprobes by a specific interaction between the protein A expressed on the S. aureus surface and the Fc fragment domain of antibodies, avoiding the use of other common methods for cell surface modifications, such as molecular covalent connection or more difficult genetic and metabolic engineering. Coupled with immunomagnetic beads, the resulting fluorescent-biotargeting bifunctional cells, i.e., biotargeting cellular beacons, can be employed as nanobioprobes for detection of viruses, bacteria, and tumor cells. With this method, H9N2 AIV can be detected specifically with a limit of 8.94 ng/mL (based on protein content). Furthermore, diverse probes for detection of different pathogens or for other biomedical applications can be easily obtained by simply changing the antibody conjugated to the cell surface.

A method based on the AFM and colloidal probe techniques was proposed to directly measure nonspecific interactions between QDs and different proteins with respective sizes and isoelectric points. Results indicated that van der Waals forces were the leading force, while electrostatic interactions also played an important role in nonspecific interactions.

Utilization of quantum dots (QDs) for single-virus tracking is highly important for understanding virus infection mechanism. However, QD labeling site of real enveloped viruses has been confined to the external envelope so far, causing the impossibility to monitor the late infection events after the loss of envelope. Herein, a strategy to label the internal nucleocapsid of enveloped virus with QDs was proposed. The nucleocapsid of enveloped baculovirus was self-biotinylated during virus replication process in host cells and subsequently labeled with streptavidin-conjugated QDs (SA-QDs). Such host cell-assisted QD labeling was proved to be reliable, specific, efficient and capable of maintaining virus infectivity. Based on such labeling, critical infection events before and after the envelope loss were monitored in real time, including single virus interacting with late endosomes and the subsequent nucleocapsid transporting into cell nucleus. Thus our established QD labeling of enveloped virus nucleocapsid with QDs enables the comprehensive single-virus tracking for deeply understanding virus infection mechanism., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Picoliter droplets were developed as microreactors for ultrafast and continuous synthesis of multi-color, water-soluble CdTe quantum dots (QDs). Through a slight change in the local controllable reaction temperature of 1-2 °C, we could obtain a series of different colored fluorescent QDs in about 1 min.

Aim: This study aimed to provide new insights into the mechanisms of hepatocellular carcinoma (HCC) invasion by simultaneously imaging tumor cells and major components of the tumor microenvironment., Materials & Methods: Formalin-fixed paraffin-embedded human HCC tissues were studied by conventional immunohistochemistry and quantum dot-based multiplexed imaging to reveal type IV collagen, LOX and tumor angiogenesis., Results: Type IV collagen degradation and repatterning in the extracellular matrix (ECM) was a continuous process, making the ECM harder, although more fragile and less resistant to cancer invasion. The distribution of LOX among cancer nests was heterogeneous, with higher expression in small cancer nests and lower expression in large cancer nests. LOX expression in cancer cells was associated with rigid stroma and tumor angiogenesis. Tumor angiogenesis occurred with type IV collagen presence. At the cancer invasion front, the ECM was hydrolyzed, with the prominent linear reorientation of type IV collagen surrounding cancer nests adjacent to neovessels., Conclusion: The visualization of the temporal-spatial relationship between type IV collagen, LOX and tumor angiogenesis revealed the coevolution process of HCC cells and their microenvironment, emphasizing an active role of the ECM during cancer invasion.

A sandwich immunoassay method for rapid detection of dual cancer biomarkers in serum on a magnetic field controllable microfluidic chip (MFCM-Chip) was established. A nickel pattern was used to generate high magnetic field gradients to increase the magnetic force on the superparamagnetic beads (SPMBs), which enabled the rapid generation of controllable SPMB patterns in microfluidic channels. The SPMB patterns could keep stable during the fast continuous washing process even at a flow rate of 50 μL/min. This approach demonstrated excellent specificity because the fast continuous washing could remove non-specifically adsorptive contaminants more efficiently than fixed volume batch washing. This approach was used to simultaneously detect carcinoma embryonic antigen (CEA) and α-fetoprotein (AFP) directly in serums. The whole on-chip detection was finished within 40 min, which was much faster than conventional enzyme-linked immunosorbent assay (ELISA) method. High luminescent streptavidin modified QDs (SA-QDs) were used as fluorescence indicators, and the detection limits were 3.5 ng/mL and 3.9 ng/mL for CEA and AFP, respectively. The linear ranges were from 10.0 ng/mL to 800.0 ng/mL. With the high sensitivity, high selectivity and short assay time, this approach could be used for rapid, high throughput detection of cancer biomarkers in clinical trials., (Copyright © 2012 Elsevier B.V. All rights reserved.)

We report a non-iterative localization algorithm that utilizes the scaling of a three-dimensional (3D) image in the axial direction and focuses on evaluating the radial symmetry center of the scaled image to achieve the desired single-particle localization. Using this approach, we analyzed simulated 3D particle images by wide-field microscopy and confocal microscopy respectively, and the 3D trajectory of quantum dots (QDs)-labeled influenza virus in live cells. Both applications indicate that the method can achieve 3D single-particle localization with a sub-pixel precision and sub-millisecond computation time. The precision is almost the same as that of the iterative nonlinear least-squares 3D Gaussian fitting method, but with two orders of magnitude higher computation speed. This approach can reduce considerably the time and costs for processing the large volume data of 3D images for 3D single-particle tracking, which is especially suited for 3D high-precision single-particle tracking, 3D single-molecule imaging and even new microscopy techniques.

Tumor microenvironment has been increasingly recognized as a complex and dynamic cancer society influencing tumor invasion and progression. The prognostic significance of this microenvironment is yet to be fully appreciated. A holistic approach to obtaining integrated information on key components in tumor microenvironment is essential. Here we reported on a quantum dots (QDs)-based simultaneous in-situ detection of infiltrating macrophages, tumor microvessels density (MVD) and neovessels maturity, in gastric cancer tissues, to obtain integrated information on these components, termed as combined tumor stromal features. These stromal features had the comparable prognostic value for overall survival, and even better prognostic value for disease-free survival, compared with traditional tumor cell-based clinico-pathological parameters. Subgroups of gastric cancer patients with favorable and unfavorable combined tumor stromal features were identified, with significantly different clinical outcomes. This study demonstrated the technical advantages of QDs-based simultaneous detection of multiple biomarkers in situ, revealed the important role of tumor stroma in cancer biology, and opened a new field to predict clinical outcome in gastric cancer from the perspectives of tumor microenvironment., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

A one-step method for synthesizing water-soluble Ag(2)S quantum dots terminated with carboxylic acid group has been reported. The crystal structure and surface of the prepared Ag(2)S quantum dots were characterized. The prepared Ag(2)S quantum dots exhibited bright photoluminescence and excellent photostabilities. The photoluminescence emissions could be tuned from visible region to near-infrared (NIR) region (from 510 nm to 1221 nm). Ultra-small sized Ag(2)S nanoclusters were synthesized with high initial monomer concentration in the current system. The in vivo imaging experiments of nude mice showed that the NIR photoluminescence of the prepared Ag(2)S quantum dots could penetrate the body of mice. Compared to the conventional NIR quantum dots, the Ag(2)S quantum dots don't contain toxic elements to body (such as Cd and Pb), thus, the prepared Ag(2)S quantum dots could serve as excellent NIR optical imaging probes and would open the opportunity to study nanodiagnostics and imaging in vivo., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Bead-based optical encoding or magnetic encoding techniques are promising in high-throughput multiplexed detection and separation of numerous species under complicated conditions. Therefore, a self-assembly strategy implemented in an organic solvent is put forward to fabricate fluorescent-magnetic dual-encoded nanospheres. Briefly, hydrophobic trioctylphosphine oxide-capped CdSe/ZnS quantum dots (QDs) and oleic acid-capped nano-γ-Fe2O3 magnetic particles are directly, selectively and controllably assembled on branched poly(ethylene imine)-coated nanospheres without any pretreatment, which is crucial to keep the high quantum yield of QDs and good dispersibility of γ-Fe2O3. Owing to the tunability of coating amounts of QDs and γ-Fe2O3 as well as controllable fluorescent emissions of deposited-QDs, dual-encoded nanospheres with different photoluminescent emissions and gradient magnetic susceptibility are constructed. Using this improved layer-by-layer self-assembly approach, deposition of hydrophobic nanoparticles onto hydrophilic carriers in organic media can be easily realized; meanwhile, fluorescent-magnetic dual-functional nanospheres can be further equipped with readable optical and magnetic addresses. The resultant fluorescent-magnetic dual-encoded nanospheres possess both the unique optical properties of QDs and the superparamagnetic properties of γ-Fe2O3, exhibiting good monodispersibility, huge encoding capacity and nanoscale particle size. Compared with the encoded microbeads reported by others, the nanometre scale of the dual-encoded nanospheres gives them minimum steric hindrance and higher flexibility.

Exploring the virus infection mechanisms is significant for defending against virus infection and providing a basis for studying endocytosis mechanisms. Single-particle tracking technique is a powerful tool to monitor virus infection in real time for obtaining dynamic information. In this study, we reported a quantum-dot-based single-particle tracking technique to efficiently and globally research the virus infection behaviors in individual cells. It was observed that many influenza viruses were moving rapidly, converging to the microtubule organizing center (MTOC), interacting with acidic endosomes, and finally entering the target endosomes for genome release, which provides a vivid portrayal of the five-stage virus infection process. This report settles a long-pending question of how viruses move and interact with acidic endosomes before genome release in the perinuclear region and also finds that influenza virus infection is likely to be a "MTOC rescue" model for genome release. The systemic technique developed in this report is expected to be widely used for studying the mechanisms of virus infection and uncovering the secrets of endocytosis., (© 2011 American Chemical Society)

A strategy is presented that involes coupling Na(2)SeO(3) reduction with the binding of silver ions and alanine in a quasi-biosystem to obtain ultrasmall, near-infrared Ag(2)Se quantum dots (QDs) with tunable fluorescence at 90 °C in aqueous solution. This strategy avoids high temperatures, high pressures, and organic solvents so that water-dispersible sub-3 nm Ag(2)Se QDs can be directly obtained. The photoluminescence of the Ag(2)Se QDs was size-dependent over a wavelength range from 700 to 820 nm, corresponding to sizes from 1.5 ± 0.4 to 2.4 ± 0.5 nm, with good monodispersity. The Ag(2)Se QDs are less cytotoxic than other nanomaterials used for similar applications. Furthermore, the NIR fluorescence of the Ag(2)Se QDs could penetrate through the abdominal cavity of a living nude mouse and could be detected on its back side, demonstrating the potential applications of these less toxic NIR Ag(2)Se QDs in bioimaging., (© 2011 American Chemical Society)

The size of C-nanodots can be electrochemically tuned by changing the applied potential during their preparation. The higher the applied potential, the smaller the resulting C-nanodots. Moreover, the surface oxidation degree of the C-nanodots can also be electrochemically tuned. The red-shift of emission independent of the size provides an insight into the luminescence mechanism of C-nanodots., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Quantum dots were proposed as new fluorochromes for use in fluorescence in-situ hybridization. EBV-encoded small RNA, the most abundant viral product in latently infected cells, was detected by quantum dot fluorescence in-situ hybridization in paraffin-embedded tissue sections of gastric carcinoma. An indirect FISH approach using quantum dots streptavidin conjugates as secondary reporters and digoxigenin labeled EBV-encoded small RNA oligonucleotide probes as detectable molecules was employed. Quantum dot fluorescence in-situ hybridization offered a slightly higher sensitivity in detecting EBV-encoded small RNA in gastric carcinoma than chromogenic in-situ hybridization. Statistical analyses showed that the detected EBV-associated gastric carcinoma was not associated with any clinicopathological parameters of the Chinese gastric carcinoma patients investigated in this study.

Wheat germ agglutinin (WGA) is a paradigm for understanding intracellular transport of lectins. As a protein exploiting the receptor-mediated endocytosis for internalization, WGA is also a valuable model system for exploring the endocytic and exocytic pathway. In this study, quantum dot-based single-particle tracking was performed to investigate the transport of WGA in live cells, revealing firstly that the endocytic and exocytic processes of WGA were both actin- and microtubule-dependent, each including five stages. The vesicle fusion event occurred near the cytomembrane, followed by two destinies with WGA: shedding to the extracellular or reversing to the cytoplasm. These findings suggest a distinct and dynamic scenario for the transport of lectins following a receptor-mediated endo/exocytic pathway in live cells. This is important for the application of lectins as drug carriers and antineoplastic drugs in medicine, and also offers insights into the pathway of endocytosis and exocytosis., (Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.)

A strategy has been put forward to fabricate PEG-interspersed nitrilotriacetic acid (NTA)-functionalized QDs by one-step self-assembly using a mixture of self-synthesized NTA-terminated amphiphilic polymer and 1,2-Distearoyl-sn-Glycero-3-Phosphoethanolamine-N-[Carboxy(Polyethylene Glycol)2000] (DSPE-PEG-COOH). The process was highly reproducible for facile functionalization of QDs via simultaneous self-assembly of biocompatible PEG molecules onto their surface. An optimized molar ratio of NTA-terminated amphiphilic polymer to DSPE-PEG-COOH was used to obtain NTA-functionalized QDs for site-specific labeling of prion proteins (PrP(C)) expressed on cell surfaces. Fabricated NTA-functionalized QDs can be a good candidate used for real-time visualization of PrP(C) in single live cells so as to clarify the nosogenesis of pathogenic scrapie prion protein (PrP(Sc))., (Copyright (c) 2010 Elsevier Ltd. All rights reserved.)

Quantum dots (QDs) are inorganic fluorescent nanocrystals with excellent properties such as tunable emission spectra and photo-bleaching resistance compared with organic dyes, which make them appropriate for applications in molecular beacons. In this work, quantum dot-based molecular beacons (QD-based MBs) were fabricated to specifically detect β-lactamase genes located in pUC18 which were responsible for antibiotic resistance in bacteria Escherichia coli (E. coli) DH5α. QD-based MBs were constructed by conjugating mercaptoacetic acid-quantum dots (MAA-QDs) with black hole quencher 2 (BHQ2) labeled thiol DNA vial metal-thiol bonds. Two types of molecular beacons, double-strands beacons and hairpin beacons, were observed in product characterization by gel electrophoresis. Using QD-based MBs, one-step FISH in tiny bacteria DH5α was realized for the first time. QD-based MBs retained their bioactivity when hybridizing with complementary target DNA, which showed excellent advantages of eliminating background noise caused by adsorption of non-specific bioprobes and achieving clearer focus of genes in plasmids pUC18, and capability of bacterial cell penetration and signal specificity in one-step in situ hybridization., (Copyright © 2010 Elsevier B.V. All rights reserved.)