Cell Bioimaging Research Articles

Quinazoli-4-one ionic liquid as a fluorescent sensor for NH3 detection: Interaction with ctDNA, theoretical investigation and live cell bioimaging

A novel quinazoli-4-one based ionic liquid, 1-(3-aminopropyl)-3-methyl-4-oxo-3,4-dihydroquinazolin-1-ium bromide (QIL) for fluorometric determination of dissolved ammonia has been successfully synthesized and characterized by spectroscopic techniques such as 1H and 13C NMR, FTIR and HRMS spectrometry. In the proposed method, QIL is converted to a fluorescent derivative by the reaction with ammonia in aqueous medium. The excitation and emission wavelengths were 250 and 436 nm, respectively. Remarkably with the reaction time of >1 s, the binding constant and detection limit was found to be 6.43 × 108 M−1 and 0.73 × 10−8 M, respectively. QIL is found to be highly selective as no interference is observed from various cations, anions, organic molecules and amino acids. The sensing mechanism was further validated by the density functional theory studies. The fluorophore exhibited great sensing property in 3.0–14.0 pH range, hence, it can be employed in diverse matrices. In addition, the fluoro-sensor is highly reversible and reusable in the presence of ctDNA molecule. Moreover, a live-cell imaging study of QIL in Drosophila larval gut tissue has also been carried out to investigate the cell permeability of QIL and its efficiency for selective detection of NH3 in cellular micro environment. To show practical applicability of the fluoro-sensor, test strip kit has been constructed. A detailed comparison table has been shown to evaluate the efficiency of this method.

Visual and quantitative monitoring of thiophenol by a novel deep-red emitting fluorescent probe in environmental and biological systems

Detection of highly toxic thiophenols in biological or environmental systems is of great importance. Therefore, fast, reliable, and sensitive probes are needed to detect thiophenols. Herein, a novel triphenylamine conjugated dicyanoisophorone-based near infrared fluorescence probe is reported to determine trace thiophenol (PhSH) levels. The probe demonstrates a distinct “turn-on” fluorescence response to thiophenol among the tested analytes and its quantum yield (Φ) increases from 0.011 to 0.142. It has low cytotoxicity with cell viability of 90–100% up to 10.0 μM of the probe, a strong anti-interference capability, a large Stokes shift (150 nm), and a fast response time (<1 min). In addition, the probe exhibits a good linear response to PhSH over the range from 0 to 15.0 μM with a detection limit of 32.3 nM (R2 = 0.9978). The detection process is also confirmed through HPLC. The practical applicability of the probe is proved by a smartphone platform, TLC kit, plant tissue imaging, soil assay, tap, and lake water analysis with good recovery values (92.3–117%), and concentration-dependent live cell bioimaging PhSH from 5.0 to 15.0 μM. Therefore, the present probe is a robust candidate for monitoring PhSH levels in biological and environmental systems.

A biphenyl thiosemicarbazide based fluorogenic chemosensor for selective recognition of Cd2+: application in cell bioimaging.

A diversified biphenyl thiosemicarbazide based chemosensor (HBMC) has been fabricated and reported for the specific detection of Cd2+ in a MeOH : H2O (4 : 1) solution. We observed a chromogenic change from colorless to light yellow colour, and it showed a "turn-on" fluorogenic change from non fluorescent to blooming cyan colour. In fluorometric titration a sharp "turn-on" emission for Cd2+ was observed with a ∼16 fold increase in fluorescence intensity value at 496 nm by incremental addition of Cd2+ ions in the MeOH : H2O (4 : 1) solution. The reversibility of the chemosensor (HBMC) was confirmed by a sequential addition of the EDTA solution. Again the binding stoichiometry of HBMC with Cd2+ was found to be 2 : 1, as confirmed by Job's plot analysis and HRMS spectra of the HBMC-Cd2+ complex. The mechanism for Cd2+ sensing in MeOH : H2O (4 : 1) is based upon the inhibition of CN isomerization and ESIPT process and simultaneously turning on the CHEF (chelation enhanced fluorescence) process. The limit of detection for Cd2+ was found to be in the order of 10-8 (M), which implies that HBMC is an efficient probe to detect Cd2+ at the microscopic level. A reusability study was performed and on-sight detection of cadmium ions by the chemosensor (HBMC) was established by dip-stick experiment. In vitro detection of Cd2+ in human breast cancer cells (MDA-MB-231) by HBMC discloses its cell permeability and biocompatible nature. Computational studies (DFT and TDDFT) with the probe HBMC and HBMC-Cd2+ complex were also performed.

Fluorescence switch based on NIR-emitting carbon dots revealing high selectivity in the rapid response and bioimaging of oxytetracycline.

The abuse of antibiotics has led to serious environmental pollution and the emergence of drug-resistant bacteria surpassing the replacement rate of antibiotics. Herein, near-infrared fluorescent carbon dots (NIR-CDs) were developed to meet the requirements for oxytetracycline (OTC) detection in food and water samples (milk, honey, and lake water) with a detection limit of 0.112 μM. These NIR-CDs, possessing excellent water-solubility, deep tissue penetration ability, and tunable optical properties, exhibit maximum emission at 790 nm (NIR-I window). Unlike traditional CDs, this novel NIR-CDs nanoprobe provides a dual response in the presence of OTC (quenching and bathochromic shifting), without obvious interference from other existing biomolecules and metal ions. Additionally, these NIR-CDs exhibit excellent photostability and multi-resistance under UV irradiation, exceptional pH stability (pH 6-12), reliable long-time exposure, and durability in ionic (NaCl) environments. Moreover, NIR-CDs and NIR-CDs@OTC are nontoxic and were successfully utilized for cell-imaging applications in normal (NIH3T3) and cancer cells (HeLa).

A novel fluorescent probe with high sensitivity for sequential detection of CN− and Al3+ in highly aqueous medium and its applications in living cell bioimaging

Benzothiazole refers to a type of heteroaromatic compound constituted by electron rich nitrogen and sulphur atoms from molecular level, and presents powerful fluorescence in solutions, resulting its widely application in fluorescent whitening agents, fluorescent dyes and functional coordination. In this paper, we focused on designing and synthesizing a benzothiazole-functionalized naphthalimide fluorescent probe NIB for sequentially detecting CN− and Al3+ in DMSO/H2O (1:9, v/v) solution using these fluorescence properties of benzothiazole. When the fluorescent probe NIB interacts with CN−, the fluorescence of the probe is quenched, indicating that NIB can specifically respond to CN−. According to the anti-interference experiment, NIB possesses strong anti-interference ability toward CN−, and its detection limit of CN− is 6.31 × 10−7 M. Interestingly, the fluorescence of the NIB-CN− system was turned back on again by adding Al3+ ions which has good anti-interference performance against other cations. The lowest detection limit of NIB-CN− system for Al3+ is 2.85 × 10−6 M. After 10 cycles, NIB still has good sequential detection performance for CN− and Al3+, which can serve as a highly sensitive logic gate for the detection of CN− and Al3+, and the “On-Off-On” switches of this fluorescence can be observed under UV light megascopically. The mechanism of sequential detection of CN− and Al3+ was further discussed by experimental and theoretical methods. The detection mechanism is not the traditional coordination competition mechanism, but the sequential addition of CN− and Al3+ changes the aggregation mode of probe molecules in the system. In addition, the potential application of this probe was fully confirmed by the preparation of probe NIB test paper and the realization of sequential detection of CN− and Al3+ in living cells (HeLa).

Artificial light-harvesting systems and their applications in photocatalysis and cell labeling

• The artificial light-harvesting systems based on non-covalent interactions are discussed. • The artificial light-harvesting systems with two-step sequential energy transfer are discussed. • The applications of artificial light-harvesting systems in photocatalysis and cell labeling are discussed. Photosynthesis is the basis for the survival of organisms in nature; consequently, the fabrication of artificial light-harvesting systems (LHSs) that simulate natural photosynthesis is of significant interest. Recently, a variety of artificial LHSs have been successfully constructed using fluorescence resonance energy transfer (FRET). However, it is crucial to fabricate artificial LHSs with a sequential energy transfer process when considering that the natural photosynthetic process involves a multistep sequential energy transfer process rather than a simple one-step energy transfer. Moreover, many previously reported LHSs have been used as imaging agents for cell labeling and bioimaging or as catalysts in photocatalytic reactions, showing promise for applications simulating natural photosynthesis. In this review, we have summarized recently published representative work on artificial LHSs. In addition, the application of LHSs in photocatalysis and cell labeling has been described in detail.

Multifunctional surface functionalized magnetic iron oxide nanoparticles for biomedical applications: A review

Magnetic iron oxide nanoparticle-based multifunctional platforms have been explored extensively in biomedical applications. Modifications and integrations of IONPs with different entities viz. organic polymer, doping with inorganic materials, loading with drug, fluorescent dye, or antibodies make them appropriate for their application in broad spectrum of biomedical fields. This review presents and summarizes the fabrication strategies of multifunctional magnetic nanoparticles based on the modification and surface functionalization of MNP. Multifunctional IONPs based recent advances covering a wide array of applications like biosensing and pathogen detection, magnetic resonance imaging (MRI) and biomarker tracking, magnetofection and gene therapy, hyperthermia and chemotherapy, drug delivery and targeted cell killing, bioimaging and therapeutics, stem cell detection and therapy, tissue engineering and organ transplant, nano-vaccines and immune system activation, microbe targeting and destruction, and COVID19 management are also covered.

Facile Synthesis of N, S-Doped Carbon Quantum Dots from Food Waste as Fluorescent Probe for Sensitive Detection of Thiamphenicol and Its Analogues in Real Food Samples along with an Application in Bioimaging.

Herein, N, S co-doped carbon quantum dots (N, S-CDs) with high absolute quantitative yield (Abs-QY) of 50.2% were produced by hydrothermal treatment of food residue crayfish shells. A new detection method of thiamphenicol (TAP) and its analogues was established by discovering the obvious fluorescence response between TAP and N, S-CDs, which achieved a wide linear range of 20–300 μg·L−1 with a detection limit (LOD) of 11.12 μg·L−1. This novel probe exhibited strong sensitivity and shows rapid response in complex food matrices (overall detection time is less than 45 min) mainly induced by static quenching. Spiked food sample recovery ranged from 97.3 to 99.34%. Further, the cell experiments of N, S-CDs were conducted, and the cell viability remained 91.76% under high concentration of N, S-CDs due to the environmentally friendly materials. The low cytotoxicity and good cytocompatibility make these N, S-CDs compatible for cell bioimaging and intracellular detection of TAP.

Bio-assembled smart nanocapsules for targeted delivery of KRAS shRNA and cancer cell bioimage

The five-year survival rate for pancreatic cancer is less than 5%. However, the current clinical multimodal therapy combined with first-line chemotherapy drugs only increases the patient's median survival from 5.0 months to 7.2 months. Consequently, a new strategy of cancer treatments is urgently needed to overcome this high-fatality disease. Through a series of biometric analyses, we found that KRAS is highly expressed in the tumor of pancreatic cancer patients, and this high expression is closely related to the poor prognosis of patients. It shows that inhibiting the expression of KRAS has great potential in gene therapy for pancreatic cancer. Given those above, we have exploited the possibility of targeted delivery of KRAS shRNA with the intelligent and bio-responsive nanomedicine to detect the special oxidative stress microenvironment of cancer cells and realize efficient cancer theranostics. Our observations demonstrate that by designing the smart self-assembled nanocapsules of melanin with fluorescent nanoclusters we can readily achieve the bio-recognition and bioimaging of cancer cells in biological solution or serum. The self-assembled nanocapsules can make a significant bio-response to the oxidative stress microenvironment of cancer cells and generate fluorescent zinc oxide Nanoclusters in situ for targeted cell bioimaging. Moreover, it can also readily facilitate cancer cell suppression through the targeted delivery of KRAS shRNA and low-temperature hyperthermia. This raises the possibility to provide a promising theranostics platform and self-assembled nanomedicine for targeted cancer diagnostics and treatments through special oxidative stress-responsive effects of cancer cells. The self-assembled nanocapsules can make a significant bio-response to the oxidative stress microenvironment of cancer cells and readily facilitate cancer cell suppression through the targeted delivery of KRAS shRNA and low-temperature hyperthermia.

Amphiphilicity-Controlled Localization of Red Emitting Bicationic Fluorophores in Tumor Cells Acting as Bio-Probes and Anticancer Drugs.

Small organic molecules arouse lively interest for their plethora of possible biological applications, such as anticancer therapy, for their ability to interact with nucleic acids, or bioimaging, thanks to their fluorescence emission. Here, a panchromatic series of styryl-azinium bicationic dyes, which have already proved to exhibit high water-solubility and significant red fluorescence in water, were investigated through spectrofluorimetric titrations to assess the extent of their association constants with DNA and RNA. Femtosecond-resolved transient absorption spectroscopy was also employed to characterize the changes in the photophysical properties of these fluorophores upon interaction with their biological targets. Finally, in vitro experiments conducted on tumor cell lines revealed that some of the bicationic fluorophores had a peculiar localization within cell nuclei exerting important antiproliferative effects, others were instead found to localize in the cytoplasm without leading to cell death, being useful to mark specific organelles in light of live cell bioimaging. Interestingly, this molecule-dependent behavior matched the different amphiphilicity featured by these bioactive compounds, which are thus expected to be caught in a tug-of-war between lipophilicity, ensured by the presence of aromatic rings and needed to pass cell membranes, and hydrophilicity, granted by charged groups and necessary for stability in aqueous media.

Microfluidic-assisted synthesis and modeling of stimuli-responsive monodispersed chitosan microgels for drug delivery applications

Droplet microfluidic has been established to synthesize and functionalize micro/nanoparticles for drug delivery and screening, biosensing, cell/tissue engineering, lab-on-a-chip, and organ-on-a-chip have attracted much attention in chemical and biomedical engineering. Chitosan (CS) has been suggested for different biomedical applications due to its unique characteristics, such as antibacterial bioactivities, immune-enhancing influences, and anticancer bioactivities. The simulation results exhibited an alternative for attaining visions in this complex method. In this regard, the role of the flow rate ratio on the CS droplet features, including the generation rate and droplet size, were thoroughly described. Based on the results, an appropriate protocol was advanced for controlling the CS droplet properties for comparing their properties, such as the rate and size of the CS droplets in the microchip. Also, a level set (LS) laminar two-phase flow system was utilized to study the CS droplet-breaking process in the Flow Focused-based microchip. The outcomes demonstrated that different sizes and geometries of CS droplets could be established via varying the several parameters that validated addressing the different challenges for several purposes like drug delivery (the droplets with smaller sizes), tissue engineering, and cell encapsulation (the droplets with larger sizes), lab-on-a-chip, organ-on-a-chip, biosensing and bioimaging (the droplets with different sizes). An experimental study was added to confirm the simulation results. A drug delivery application was established to verify the claim.

Carboxymethylcellulose biofunctionalized ternary quantum dots for subcellular-targeted brain cancer nanotheranostics

Among the most lethal forms of cancer, malignant brain tumors persist as one of the greatest challenges faced by oncologists, where nanotechnology-driven theranostics can play a critical role in developing novel polymer-based supramolecular nanoarchitectures with multifunctional and multi-modal characteristics to fight cancer. However, it is virtually a consensus that, besides the complexity of active delivering anticancer drugs by the nanocarriers to the tumor site, the current evaluation methods primarily relying on in vitro assays and in vivo animal models have been accounted for the low translational effectiveness to clinical applications. In this view, the chick chorioallantoic membrane (CAM) assay has been increasingly recognized as one of the best preclinical models to study the effects of anticancer drugs on the tumor microenvironment (TME). Thus, in this study, we designed, characterized, and developed novel hybrid nanostructures encompassing chemically functionalized carboxymethylcellulose (CMC) with mitochondria-targeting pro-apoptotic peptide (KLA) and cell-penetrating moiety (cysteine, CYS) with fluorescent inorganic semiconductor (Ag-In-S, AIS) for simultaneously bioimaging and inducing glioblastoma cancer cell (U-87 MG, GBM) death. The results demonstrated that the CMC-peptide macromolecules produced supramolecular vesicle-like nanostructures with aqueous colloidal stability suitable as nanocarriers for passive and active targeting of cancer tumors. The optical properties and physicochemical features of the nanoconjugates confirmed their suitability as photoluminescent nanoprobes for cell bioimaging and intracellular tracking. Moreover, the results in vitro demonstrated a notable killing activity towards GBM cells of cysteine-bearing CMC conjugates coupled with pro-apoptotic KLA peptides. More importantly, compared to doxorubicin (DOX), a model anticancer drug in chemotherapy that is highly toxic, these innovative nanohybrids nanoconjugates displayed higher lethality against U-87 MG cancer cells. In vivo CAM assays validated these findings where the nanohybrids demonstrated a significant reduction of GBM tumor progression (41% area) and evidenced an antiangiogenic activity. These results pave the way for developing polymer-based hybrid nanoarchitectonics applied as targeted multifunctional theranostics for simultaneous imaging and therapy against glioblastoma while possibly reducing the systemic toxicity and side-effects of conventional anticancer chemotherapeutic agents.

Research update of emergent gold nanoclusters: A reinforced approach towards evolution, synthesis mechanism and application.

Fluorescent biosensors and imaging devices have gained fervent consideration due to their prime functionality in biological. Among fluorescent nanomaterial (FNMs), the ultra-small gold nanoclusters (AuNCs) have gained promising attention with respect to extra-ordinary properties of bright fluorescence, economical synthesis, higher photostability, and biocompatibility, and deep tissue penetration. Therefore, the prior decades comprehended the revolutions in the field of real-time monitoring devices, nanotechnology-based biosensing, and bioimaging sensors. The present review primarily focuses on metal NCs (MNCs) and their advantages, a brief introduction of AuNCs along with history prospective and development, fundamental aspects regarding AuNCs quality and fluorescence, limitations and advantages of AuNCs, sensing mechanism, expected synthesis principle, and summarized the recent progress of AuNCs probe over the last 3 years (2019-2021) with their respective detection and bioimaging mechanism and synthesis principle. Moreover, the present work also serves as a novel stratagem for the preparation and potential applications of multifunctional AuNCs nano-systems responsiveness for various analytes detection and cell bioimaging with respective examples. At last, we described the challenges associated with the application of AuNCs based on recent signs of progress.

Recent advances in non-plasmonic surface-enhanced Raman spectroscopy nanostructures for biomedical applications.

Surface-enhanced Raman spectroscopy (SERS) is an emerging powerful vibrational technique offering unprecedented opportunities in biomedical science for the sensitive detection of biomarkers and the imaging and tracking of biological samples. Conventional SERS detection is based on the use of plasmonic substrates (e.g., Au and Ag nanostructures), which exhibit very high enhancement factors (EF=1010 -1011 ) but suffers from serious limitations, including light-induced local heating effect due to ohmic loss and expensive price. These drawbacks may limit detection accuracy and large-scaled practical applications. In this review, we focus on alternative approaches based on plasmon-free SERS detection on low-cost nanostructures, such as carbons, oxides, chalcogenides, polymers, silicons, and so forth. The mechanism of non-plasmonic SERS detection has been attributed to interfacial charge transfer between the substrate and the adsorbed molecules, with no photothermal side-effects but usually less EF compared with plasmonic nanostructures. The strategies to improve Raman signal detection, through the tailoring of substrate composition, structure, and surface chemistry, is reviewed and discussed. The biomedical applications, for example, SERS cell characterization, biosensing, and bioimaging are also presented, highlighting the importance of substrate surface functionalization to achieve sensitive, accurate analysis, and excellent biocompatibility. This article is categorized under: Diagnostic Tools > Diagnostic Nanodevices Diagnostic Tools > Biosensing Diagnostic Tools > In Vivo Nanodiagnostics and Imaging.

Phosphorogenic Iridium(III) bis‐Tetrazine Complexes for Bioorthogonal Peptide Stapling, Bioimaging, Photocytotoxic Applications, and the Construction of Nanosized Hydrogels

AbstractThe dual functionality of 1,2,4,5‐tetrazine as a bioorthogonal reactive unit and a luminescence quencher has shaped tetrazine‐based probes as attractive candidates for luminogenic labeling of biomolecules in living systems. In this work, three cyclometalated iridium(III) complexes featuring two tetrazine units were synthesized and characterized. Upon photoexcitation, the complexes were non‐emissive but displayed up to 3900‐fold emission enhancement upon the inverse electron‐demand Diels–Alder (IEDDA) [4+2] cycloaddition with (1R,8S,9s)‐bicyclo[6.1.0]non‐4‐yne (BCN) substrates. The rapid reaction kinetics (k2 up to 1.47×104 M−1 s−1) of the complexes toward BCN substrates allowed effective peptide labeling. The complexes were also applied as live cell bioimaging reagents and photocytotoxic agents. One of the complexes was utilized in the preparation of luminescent nanosized hydrogels that exhibited interesting cargo delivery properties.

Phosphorogenic Iridium(III) bis-Tetrazine Complexes for Bioorthogonal Peptide Stapling, Bioimaging, Photocytotoxic Applications, and the Construction of Nanosized Hydrogels.

The dual functionality of 1,2,4,5-tetrazine as a bioorthogonal reactive unit and a luminescence quencher has shaped tetrazine-based probes as attractive candidates for luminogenic labeling of biomolecules in living systems. In this work, three cyclometalated iridium(III) complexes featuring two tetrazine units were synthesized and characterized. Upon photoexcitation, the complexes were non-emissive but displayed up to 3900-fold emission enhancement upon the inverse electron-demand Diels-Alder (IEDDA) [4+2] cycloaddition with (1R,8S,9s)-bicyclo[6.1.0]non-4-yne (BCN) substrates. The rapid reaction kinetics (k2 up to 1.47×104 M-1 s-1 ) of the complexes toward BCN substrates allowed effective peptide labeling. The complexes were also applied as live cell bioimaging reagents and photocytotoxic agents. One of the complexes was utilized in the preparation of luminescent nanosized hydrogels that exhibited interesting cargo delivery properties.

An unconventional nano-AIEgen originating from a natural plant polyphenol for multicolor bioimaging

Biocompatible aggregation-induced emission (AIE) materials from natural resources are of great interest for a variety of potential applications. Here, we report the unconventional AIE nature of the green tea polyphenol epigallocatechin gallate (EGCG), which is investigated by observing the disappearance and reoccurrence of fluorescence when EGCG is dissolved and recrystallized in aqueous solutions. The intermolecular through-space conjugation and structural rigidification induced by multiple intermolecular H-bonds play critical roles in the AIE phenomenon. This inspires the development of a multicolored, monodisperse, photostable, and non-toxic nano-AIE luminogen (nano-AIEgen), which is simply prepared by polyphenol-amine-based crosslinking in aqueous solutions, demonstrating great potential for living cell bioimaging. Our strategy to develop nano-AIEgens using AIE-active hydrophilic natural products—e.g., plant polyphenols, such as tannic acid, that exhibit similar AIE characteristics—can help push the boundary of the exploration of various novel, large-scale, biocompatible, water-soluble, and degradable AIE materials from natural resources. • AIE phenomenon for EGCG in the solid state • The AIE activity results from through-space conjugation by intermolecular H-bonds • The EGCG nano-AIEgens may be successfully used to image living cells As a plant polyphenol, epigallocatechin gallate is highly soluble in water and non-toxic. Here, Lu et al. demonstrate an interesting AIE phenomenon in common natural products and propose strategies for screening and developing hydrophilic AIEgens from natural products.

Multiple heteroatom-doped photoluminescent carbon dots for ratiometric detection of Hg2+ ions in cell imaging and environmental applications.

Photoluminescence detection and imaging of Hg2+ ions in the biochemical living system are of great importance. In this study, a new photoluminescent probe based on nitrogen (N), sulfur (S), and boron (B) multiple heteroatom co-doped carbon dots (NSB-CDs) is synthesized for the ratiometric detection of Hg2+ ions. The prepared NSB-CDs possess good aqueous solubility, excellent pH and ionic stability, excitation dependency, and high quantum yield (QY = 17.6%). The ratiometric photoluminescent sensor NSB-CDs exhibit high selectivity, sensitivity, and interference towards Hg2+ ions over other metal ions. After adding Hg2+ ions, the emission intensity of the NSB-CDs exhibits a large redshift from 452 to 496 nm (up to 44 nm), corresponding to a notable change from blue to green emission in aqueous solutions. The association constant (Ka), the limit of detection (LOD), and the limit of quantification (LOQ) for NSB-CDs/Hg2+ complex are calculated to be 3.6 × 104 M-1, 3.1 × 10-9 M, and 10.4 × 10-9 M, respectively, in the range of 0-30 × 10-6 M. The live cell bioimaging of HCT-116 cells with NSB-CDs validates the application of multicolor imaging for the detection of Hg2+ ions in aqueous media and biological systems. Moreover, the potential use of the NSB-CDs/Hg2+ complex for real sample analysis is demonstrated.

Dual-Locked Near-Infrared Fluorescent Probes for Precise Detection of Melanoma via Hydrogen Peroxide-Tyrosinase Cascade Activation.

Activity-based near-infrared (NIR) fluorescent probes provide powerful tools for diagnosis of diseases. However, most of these probes suffer from low specificity due to "off-target" reaction. The dual-locked strategy, which utilizes two biomarkers as triggers, can increase the specificity and precision of diagnosis. Here, we report a dual-locked NIR probe, MB-m-borate, which releases fluorophore methylene blue (MB) after hydrogen peroxide-tyrosinase (H2O2-TYR) cascade activation. Both MB-m-borate and its intermediate MB-m-phenol (the product after H2O2 activation) show almost nondetectable fluorescence. MB-m-borate exhibits "turn on" fluorescence upon H2O2-TYR cascade activation. The further live cell bioimaging results indicate that MB-m-borate only responds to melanoma cells, providing it as a robust probe for precise detection of melanoma. Finally, the probe is applied for the diagnosis of melanoma in vivo with a xenogeneic mouse model.

Multimodal on-chip nanoscopy and quantitative phase imaging reveals the nanoscale morphology of liver sinusoidal endothelial cells

Visualization of three-dimensional (3D) morphological changes in the subcellular structures of a biological specimen is a major challenge in life science. Here, we present an integrated chip-based optical nanoscopy combined with quantitative phase microscopy (QPM) to obtain 3D morphology of liver sinusoidal endothelial cells (LSEC). LSEC have unique morphology with small nanopores (50-300 nm in diameter) in the plasma membrane, called fenestrations. The fenestrations are grouped in discrete clusters, which are around 100 to 200 nm thick. Thus, imaging and quantification of fenestrations and sieve plate thickness require resolution and sensitivity of sub-100 nm along both the lateral and the axial directions, respectively. In chip-based nanoscopy, the optical waveguides are used both for hosting and illuminating the sample. The fluorescence signal is captured by an upright microscope, which is converted into a Linnik-type interferometer to sequentially acquire both superresolved images and phase information of the sample. The multimodal microscope provided an estimate of the fenestration diameter of 119 ± 53 nm and average thickness of the sieve plates of 136.6 ± 42.4 nm, assuming the constant refractive index of cell membrane to be 1.38. Further, LSEC were treated with cytochalasin B to demonstrate the possibility of precise detection in the cell height. The mean phase value of the fenestrated area in normal and treated cells was found to be 161 ± 50 mrad and 109 ± 49 mrad, respectively. The proposed multimodal technique offers nanoscale visualization of both the lateral size and the thickness map, which would be of broader interest in the fields of cell biology and bioimaging.