Increase In Fluorescence Research Articles

Detection of Lectin Clustering in Self‐Assembled, Glycan‐Functionalized Amphiphiles by Aggregation‐Induced Emission Luminophores

AbstractAmphiphilic glycan‐functionalized oligomers are derived by solid‐phase polymer synthesis and applied in both, self‐assembled micelles as well as giant unilamellar vesicles, as simplified models of the cell's glycocalyx. Additionally, an aggregation‐induced luminophore is introduced into the amphiphilic glycomacromolecules showing no fluorescence when the molecule is free in solution. Combining glycomacromolecules carrying a binding glycan motif and the luminophore with glycomacromolecules or other amphiphiles with no binding motifs and no luminophore in self‐assembled structures, micelles and vesicles exhibiting no or only very little fluorescence are obtained. Only upon clustering of the binding glycan motifs through interaction with a multivalent lectin receptor, an increase in fluorescence is observed. Thus, clustering events within these self‐assembled structures can be detected and localized.

Abstract 10611: Inhibition of Sars-cov-2 Viral Channel Activity Using FDA-Approved Channel Modulators Independent of Variants

Introduction: SARS-CoV-2 has undergone mutations, yielding clinically relevant variants such as Delta and Omicron. Early detection of virus and effective therapeutic treatment independent of variants help control virus spread. Hypothesis: We hypothesized that in the genome of SARS-CoV-2 two highly conserved viroporins Orf3a and E channels directly related to virus replication may be explored as a target for detection and inhibition of viral replication independent of variants using FDA-approved ion channel modulators. Methods: A combination of a fluorescence potassium ion assay with three channel modulators (4-aminopyridine, emodin-Orf3a channel blocker, gliclazide-E channel blocker) in a 96-well plate format was developed to detect SARS-CoV-2 Orf3a/E channel activity. We subtracted the fluorescence signals in the absence and presence of emodin/gliclazide to detect Orf3a and E channel activity. Two FDA-approved drugs, amantadine (an antiviral) and amitriptyline (an antidepressant) that are also ion channel blockers, were used to test whether they can inhibit Orf3a/E channel activity in isolated virus (UV inactivated) and in nasal swab samples from Covid-19 patients. Twenty-one (21) Delta and thirty (30) Omicron nasal swab samples were used. Variants were confirmed by PCR sequencing. Results: Orf3a and E form non-voltage-gated ion channels that are highly conserved in SARS-CoV-2 variants. In UV-inactivated SARS-CoV-2 Alpha, Beta, and Delta variants, significant channel activity of Orf3a/E was detected based on an increase in fluorescence induced by 4-aminopyridine, and this increase in fluorescence was inhibited by emodin and gliclazide (IC 50 = 0.42mM). Amantadine and amitriptyline can inhibit Delta virus with amitriptyline being a more potent inhibitor than amantadine. In swab samples of Delta variants, both amitriptyline and amantadine inhibited the channel activity of viral proteins with IC 50 values of 0.73mM and 1.11mM, respectively. In swab samples of Omicron variants, amitriptyline inhibited the channel activity with an IC 50 of 0.76mM. Conclusions: We developed an efficient method to screen FDA-approved ion channel modulators that can be repurposed to inhibit SARS-CoV-2 viral replication independent of variants.

Abstract 12752: Ultrasound-Responsive Liposome Enhanced Gene Delivery for Heart Disease

Introduction: Genome editor-CRISPR/Cas9 holds much promise for treatment of heart disease. A critical step is the development of gene transfer platform that can deliver genes into heart muscle efficiently. We have developed ultrasound (US)-responsive echogenic liposomes (ELIP) for controlled payload release, and enhanced payload penetration upon ultrasound application. Hypothesis: We hypothesize that ELIP in combination with ultrasound application can enhance gene delivery in cardiomyocytes. Methods: FITC (green fluorescence) labeled NF-kB oligonucleotides were used as a prototype for gene delivery. ELIP containing NF-kB-FITC (ELIP-NF-kB-FITC) were prepared by a conventional procedure of hydrating the lipid film, sonication, freezing and drying. Rats were divided into control, ELIP-NF-kB-FITC without US, and ELIP-NF-kB-FITC with US. ELIP-NF-kB-FITC were administered into the left ventricle by injection with the aorta clamped. Ultrasound (1 MHz at 0.5 W/cm2, 100% duty cycle) was applied to the heart for 5 min. Thereafter, heart was excise, fixed and imaged by fluorescent microscopy. The fluorescent intensity of the left ventricular wall was quantified by NIH ImageJ software. Results: ELIP-NF-kB-FITC delivered without ultrasound application showed limited green fluorescence in the heart. Ultrasound (US) in combination with ELIP enhanced NF-kB-FITC penetration into the ventricular walls (Fig. 1A). Quantitatively, ultrasound provided 2-fold increase (2.0 ± 0.27, P < 0.01 vs control) in the mean green fluorescence (Fig. 1 B). Conclusions: Ultrasound application in combination with ELIP enhanced gene delivery into the ventricular walls. This platform offers a promising delivery system of CRISPR components for the correction of cardiomyopathies caused by gene mutation/change.

Reactive Detection of Hydrazine by 6‐Hydroxy‐2‐Naphthonitrile Levulinate Anchored Colorimetric and Fluorescent Probe

AbstractHerein, a new reaction‐based fluorescent probe 6‐cyanonaphthalen‐2‐yl 4‐oxopentanoate (NPCN‐OLV) was designed and synthesized for the detection of hydrazine (N2H4). The addition of N2H4 to a solution of probe NPCN‐OLV resulted in a striking fluorescence increase, moreover, it is complemented by a visual color change from colorless to yellow. Importantly, this distinct color response indicates that probe NPCN‐OLV could be implemented as a colorimetric tool for the recognition of N2H4. N2H4 can be detected quantitatively. The lower detection limit for the N2H4 is 5 μM where actual fluorescence observed and gradually increased with concentration. The probe NPCN‐OLV served the selective visible naked‐eye sensing of hydrazine in the presence of representative metal ions and common anions. Along with that we have also demonstrated the use of paper strip for the qualitative detection of hydrazine, which showed an evident color change (sky blue) only with the paper strip with hydrazine in PBS buffer.

The physiological and phenotypic response of Salvia aethiopis L. to herbicides

Salvia aethiopis is an aromatic species within the Lamiaceae family, used both as a herb and as an ornamental plant, the yield and quality of which, when cultivated on a commercial basis, is frequently affected by weed infestation. Therefore, the application of herbicide is often required for the commercial production of this species. Considering which, for the current study, a pot experiment based upon a completely randomized design, with three replications, was conducted to evaluate the physiological and growth response of S. aethiopis to five commonly used herbicides (bentazon, phenmedipham/desmedipham, metribuzin, oxyfluorfen and oxadiargyl). These herbicides were applied at three concentrations (0.75X, 1X and 1.25X where X is equal to the recommended dose of herbicide). The data obtained from this study, demonstrated that herbicide application led to an increase in crop damage, minimum fluorescence (F0) and malondialdehyde content (MDA) and a concomitant decrease in the maximum quantum efficiency of PSΙΙ (FV/FM), maximum fluorescence (FM), SPAD value, relative water content (RWC), plant height and shoot and root dry weight as compared to the control. However, the level of response/tolerance was dependent upon the specific herbicide and the dose applied. Of the selected herbicides, S.aethiopis has shown to display the most tolerance to bentazon. As such, these data indicate that of the herbicides tested, bentazon should be the preferred herbicide used for broad leafed weed control, during the commercial production of this species.

Development of a Rapid and Sensitive Fluorescence Sensing Method for the Detection of Acetaldehyde in Alcoholic Beverages.

Acetaldehyde is regarded as an important flavor compound in alcoholic beverages. With the advantages of rapidity, low cost and high sensitivity, fluorescent probe could be used as a new tool for the detection of acetaldehyde. Here, an effective fluorescence sensing method based on fluorescent probe N1 (FPN1) was established in this study. The function of FPN1 relies on the nucleophile substitution reaction and photoinduced electron transfer (PET), resulting in a fluorescence increase. Remarkably, the pretreatment background removal method (BRM) was successfully applied for removal of the interference of pyruvate and acetal. The linearity range (LR), limit of detection (LOD) and recovery of the fluorescence sensing method with BRM were 0.0053-200 mg/L, 0.0016 mg/L and 94.02-108.12%, respectively, which showed a broader detection range and better performance on sensitivity compared with the traditional quantitation using gas chromatography (GC). Furthermore, successful application of the method in real samples indicated the advantages of low-cost and rapidity for small-scale detection while assuring the accuracy, which provides a new strategy for the detection of acetaldehyde concentration in alcoholic beverages.

Selective turn-on fluorescence detection of formaldehyde in the gas phase

Reliable detection of formaldehyde in gas phase still remains a challenge regarding either sensitivity or selectivity or both. In this study we report on a simple, unique sensor composite, which enables efficient detection of gaseous formaldehyde with both low limit of detection (LOD) and extremely high selectivity. The primary components of the sensor composite include a hydroxylamine sulfuric acid salt ((NH2OH)2·H2SO4) and a fluorescence turn-on probe, which both are coated within the porous matrix of a cellulose plate. The fluorescence probe is based on a perylene diimide (PDI) molecule modified with an aniline group, which is non-fluorescent in its pristine state due to the photoinduced intramolecular electron transfer. Upon interaction with formaldehyde, the hydroxylamine salt will be converted to the corresponding Schiff base through aldimine condensation, releasing the sulfuric acid. The acid released will protonate the aniline group on PDI, resulting in a quick fluorescence turn-on. By measuring the fluorescence increase we can detect formaldehyde both quantitatively and specifically. Under the optimized condition, the sensor demonstrated a LOD of 3.7 ppb, which is much lower than the air quality threshold set by WHO. The sensor was also proven highly selective towards formaldehyde, with negligible response towards other common chemicals. The work reported will not only provide a new sensor for formaldehyde, but moreover open a unique design pathway for sensor composite that combines reactive components with various fluorescence indicators so as to enable highly efficient detection of many other air pollutants and toxic chemicals.

Polarity-Dependent Twisted Intramolecular Charge Transfer in Diethylamino Coumarin Revealed by Ultrafast Spectroscopy

Twisting intramolecular charge transfer (TICT) is a common nonradiative relaxation pathway for a molecule with a flexible substituent, effectively reducing the fluorescence quantum yield (FQY) by swift twisting motions. In this work, we investigate coumarin 481 (C481) that contains a diethylamino group in solution by femtosecond transient absorption (fs-TA), femtosecond stimulated Raman spectroscopy (FSRS), and theoretical calculations, aided by coumarin 153 with conformational locking of the alkyl arms as a control sample. In different solvents with decreasing polarity, the transition energy barrier between the fluorescent state and TICT state increases, leading to an increase of the FQY. Correlating the fluorescence decay time constant with solvent polarity and viscosity parameters, the multivariable linear regression analysis indicates that the chromophore’s nonradiative relaxation pathway is affected by both hydrogen (H)-bond donating and accepting capabilities as well as dipolarity of the solvent. Results from the ground- and excited-state FSRS shed important light on structural dynamics of C481 undergoing prompt light-induced intramolecular charge transfer from the diethylamino group toward –C=O and –CF3 groups, while the excited-state C=O stretch marker band tracks initial solvation and vibrational cooling dynamics in aprotic and protic solvents (regardless of polarity) as well as H-bonding dynamics in the fluorescent state for C481 in high-polarity protic solvents like methanol. The uncovered mechanistic insights into the molecular origin for the fluorogenicity of C481 as an environment-polarity sensor substantiate the generality of ultrafast TICT state formation of flexible molecules in solution, and the site-dependent substituent(s) as an effective route to modulate the fluorescence properties for such compact, engineerable, and versatile chemosensors.

Chloroquine disrupts zinc storage granules in primary Malpighian tubule cells of Drosophila melanogaster.

Contrasting reports exist in the literature regarding the effect of chloroquine treatment on cellular zinc uptake or secretion. Here, we tested the effect of chloroquine administration in the Drosophila model organism. We show that larvae grown on a diet supplemented with 2.5mg/ml chloroquine lose up to 50% of their stored zinc and around 10% of their total potassium content. This defect in chloroquine-treated animals correlates with the appearance of abnormal autophagolysosomes in the principal cells of the Malpighian tubules, where zinc storage granules reside. We further show that the reported increase of Fluozin-3 fluorescence following treatment of cells with 300 μM chloroquine for 1 h may not reflect increased zinc accumulation, since a similar treatment in Madin-Darby canine kidney cells results in a 36% decrease in their total zinc content. Thus, chloroquine should not be considered a zinc ionophore. Zinc supplementation plus chloroquine treatment restored zinc content both in vivo and in vitro, without correcting autophagic or other ionic alterations, notably in potassium, associated with the chloroquine treatment. We suggest that chloroquine or hydroxychloroquine administration to patients could reduce intracellular zinc storage pools and be part of the drug's mechanism of action.

Developing a fluorometric urease activity microplate assay suitable for automated microbioreactor experiments.

Quantifying urease activity is an important task for Microbial Induced Calcite Precipitation research. A new urease activity microplate assay using a fluorescent pH indicator is presented. The method is also suitable for automated measurements during microbioreactor experiments. The assay reagent consists of the green fluorescent pH-indicator fluorescein, urea and a phosphate buffer. After sample addition, the microbial urease hydrolyses urea, which results in a pH and hence fluorescence increase. The fluorescence signal can be measured with a microplate reader or with the microbioreactor system BioLector, allowing for automated urease activity measurements during cultivation experiments. In both measurement systems, the fluorescence signal slope highly correlates with the urease activity measured offline with standard methods. Automated measurement is possible, as no sample preparation such as centrifugation or adjusting of the optical density is required. The assay was developed so that the culture samples turbidity, salinity or buffer concentration does not have a negative impact on the fluorescence signal. The assay allows for straightforward, non-hazardous, parallelized, cheap and reliable measurements, making research on ureolytic bacteria for Microbial Induced Calcite Precipitation more efficient. The assay could be adapted to other enzymes, which have a strong impact on the pH value.

A high-performance genetically encoded fluorescent indicator for in vivo cAMP imaging

cAMP is a key second messenger that regulates diverse cellular functions including neural plasticity. However, the spatiotemporal dynamics of intracellular cAMP in intact organisms are largely unknown due to low sensitivity and/or brightness of current genetically encoded fluorescent cAMP indicators. Here, we report the development of the new circularly permuted GFP (cpGFP)-based cAMP indicator G-Flamp1, which exhibits a large fluorescence increase (a maximum ΔF/F0 of 1100% in HEK293T cells), decent brightness, appropriate affinity (a Kd of 2.17 μM) and fast response kinetics (an association and dissociation half-time of 0.20 and 0.087 s, respectively). Furthermore, the crystal structure of the cAMP-bound G-Flamp1 reveals one linker connecting the cAMP-binding domain to cpGFP adopts a distorted β-strand conformation that may serve as a fluorescence modulation switch. We demonstrate that G-Flamp1 enables sensitive monitoring of endogenous cAMP signals in brain regions that are implicated in learning and motor control in living organisms such as fruit flies and mice.

Synthesis and application of a novel fluorescent probe for detection of hydrogen sulfide in real samples

ABSTRACT A novel reactive fluorescent hydrogen sulphide (H2S) probe (T-DNBS) consisting of 2,4-dinitrobenzenesulfonyl (DNBS) units has been synthesised from 4,7-dibromo-2,1,3-benzothidiazole. Adding H2S to probe T-DNBS solution resulted in a marked fluorescence increase accompanied by a visual colour change from colourless to pale red. Probe T-DNBS performed excellent sensitivity and selectivity for H2S, substantial fluorescence enhancement, can be detected quantitatively in the concentration range of 0–125 μM and the low detection limit was 77.78 nM. In addition, the probe T-DNBS can track H2S in red wine samples with high accuracy, high recovery and practical value in food analysis.

Photophysical characterization of coumarin and rhodanine derivatives as viscosity sensitive fluorescence turn-on probes

In this study, coumarin and benzylidene rhodanine derivatives, Cou-A and Rho-A, were synthesized and characterized as viscosity-sensitive molecular platforms. Cou-A and Rho-A showed quenched fluorescence behaviour in low viscosity polar solvents. However, when the solvent viscosity was increased from 2.5 cP to 1150 cP using water-glycerol mixtures, both probes displayed fluorescence turn-on responses. Cou-A showed a 27-fold increase in fluorescence at 476 nm, while Rho-A showed a 508-fold increase in fluorescence at 575 nm. Both Cou-A and Rho-A showed a linear relationship between fluorescence intensity (log I) and solvent viscosity (log η), with viscosity sensitivity values of 0.554 and 0.996, respectively. The reported probes' fluorescence profiles were unaffected by the presence of metal ions or the pH of the solvent medium. Further studies suggest that the fluorescence quantum yield of these probes is dependent on both viscosity and temperature.

Bioorthogonal Lanthanide Molecular Probes for Near-Infrared Fluorescence and Mass Spectrometry Imaging.

We report here the development of clickable and highly near-infrared (NIR) fluorescent lanthanide (Ln) complexes for bioorthogonal labeling of biomolecules. These azide- or alkyne-functionalized Ln complexes are hydrophilic and fluorogenic, exhibiting a strong increase of NIR fluorescence upon conjugation with biomolecules. Metabolic labeling of biomolecules with azide or alkyne, followed by click labeling with the Ln complexes, enables NIR fluorescence (NIRF) imaging of DNA, RNA, proteins, and glycans in cells. Furthermore, multicolor imaging is performed by combining click-labeling with the Ln complexes and immunostaining. In addition, the Ln complexes is compatible with click-expansion microscopy (click-ExM), which enables high-resolution NIRF imaging of cellular glycoproteins. Finally, the Ln complexes can be used for time-of-flight secondary-ion mass spectrometry (ToF-SIMS) imaging, thus achieving the first example of dual-modal imaging combining NIRF and SIMS microscopies.

Bioorthogonal Lanthanide Molecular Probes for Near‐Infrared Fluorescence and Mass Spectrometry Imaging

AbstractWe report here the development of clickable and highly near‐infrared (NIR) fluorescent lanthanide (Ln) complexes for bioorthogonal labeling of biomolecules. These azide‐ or alkyne‐functionalized Ln complexes are hydrophilic and fluorogenic, exhibiting a strong increase of NIR fluorescence upon conjugation with biomolecules. Metabolic labeling of biomolecules with azide or alkyne, followed by click labeling with the Ln complexes, enables NIR fluorescence (NIRF) imaging of DNA, RNA, proteins, and glycans in cells. Furthermore, multicolor imaging is performed by combining click‐labeling with the Ln complexes and immunostaining. In addition, the Ln complexes is compatible with click‐expansion microscopy (click‐ExM), which enables high‐resolution NIRF imaging of cellular glycoproteins. Finally, the Ln complexes can be used for time‐of‐flight secondary‐ion mass spectrometry (ToF‐SIMS) imaging, thus achieving the first example of dual‐modal imaging combining NIRF and SIMS microscopies.

Effects of ultrasonic treatment on ovomucin: Structure, functional properties and bioactivity

The effects of ultrasonic treatment on the structure, functional properties and bioactivity of Ovomucin (OVM) were investigated in this study. Ultrasonic treatment could significantly enhance OVM solubility without destroying protein molecules. The secondary structure changes, including β-sheet reduction and random coil increase, indicate more disorder in OVM structure. After ultrasonic treatment, the OVM molecule was unfolded partially, resulting in the exposure of hydrophobic regions. The changes in OVM molecules led to an increase in intrinsic fluorescence and surface hydrophobicity. By detecting the particle size of protein solution, it was confirmed that ultrasonic treatment disassembled the OVM aggregations causing a smaller particle size. Field emission scanning electron microscopy (FE-SEM) images showed that ultrasonic cavitation significantly reduced the tendency of OVM to form stacked lamellar structure. Those changes in structure resulted in the improvement of foaming, emulsification and antioxidant capacity of OVM. Meanwhile, the detection results of ELISA showed that ultrasonic treatment did not change the biological activity of OVM. These results suggested that the relatively gentle ultrasound treatment could be utilized as a potential approach to modify OVM for property improvement.

A novel H2O2-activated NIR fluorescent probe for imaging β-amyloid fibrils and mitochondrial viscosity

Alzheimer's disease (AD) is an incurable chronic neurodegenerative disease. Until now, β-amyloid, mitochondrial viscosity and the relatively high concentration of hydrogen peroxide (H2O2) have been reported to be related with AD undergoing mutual interactions. In this work, a novel NIR fluorescent probe CBP was designed, synthesized and could be activated by H2O2 to release CBD, which was used to image β-amyloid fibrils in brain slices from transgenic mouse model and mitochondrial viscosity in living cells. CBP itself showed weak response towards β-amyloid and viscosity. However, upon exposure to H2O2, CBP was oxidized to the product CBD, which was accompanied with a slight fluorescence “turn-on” effect, attributing to the departure of the quenching group of the boronic acid pinacol ester. The resulting product CBP under H2O2 was highly responsive to the viscosity increase with a remarkable fluorescence enhancement at 635 nm by 53.5 folds, and simultaneously exhibited high specificity and selectivity towards Aβ1-42 aggregates with a fluorescence increase at 618 nm. Furthermore, CBP under H2O2 was applied in imaging β-amyloid fibrils in brain slices of AD transgenic mice and mitochondrial viscosity at the cellar level. Therefore, this novel NIR H2O2-activated fluorescent probe might serve as a dual imaging agent for β-amyloid fibrils and mitochondrial viscosity with low background, attributing to the H2O2-activation-induced fluorescence enhancements.

The intriguing dose-dependent effect of selected amphiphilic compounds on insulin amyloid aggregation: Focus on a cholesterol-based detergent, Chobimalt.

The amyloidogenic self-assembly of many peptides and proteins largely depends on external conditions. Among amyloid-prone proteins, insulin attracts attention because of its physiological and therapeutic importance. In the present work, the amyloid aggregation of insulin is studied in the presence of cholesterol-based detergent, Chobimalt. The strategy to elucidate the Chobimalt-induced effect on insulin fibrillogenesis is based on performing the concentration- and time-dependent analysis using a combination of different experimental techniques, such as ThT fluorescence assay, CD, AFM, SANS, and SAXS. While at the lowest Chobimalt concentration (0.1 µM; insulin to Chobimalt molar ratio of 1:0.004) the formation of insulin fibrils was not affected, the gradual increase of Chobimalt concentration (up to 100 µM; molar ratio of 1:4) led to a significant increase in ThT fluorescence, and the maximal ThT fluorescence was 3-4-fold higher than the control insulin fibril’s ThT fluorescence intensity. Kinetic studies confirm the dose-dependent experimental results. Depending on the concentration of Chobimalt, either (i) no effect is observed, or (ii) significantly, ∼10-times prolonged lag-phases accompanied by the substantial, ∼ 3-fold higher relative ThT fluorescence intensities at the steady-state phase are recorded. In addition, at certain concentrations of Chobimalt, changes in the elongation-phase are noticed. An increase in the Chobimalt concentrations also triggers the formation of insulin fibrils with sharply altered morphological appearance. The fibrils appear to be more flexible and wavy-like with a tendency to form circles. SANS and SAXS data also revealed the morphology changes of amyloid fibrils in the presence of Chobimalt. Amyloid aggregation requires the formation of unfolded intermediates, which subsequently generate amyloidogenic nuclei. We hypothesize that the different morphology of the formed insulin fibrils is the result of the gradual binding of Chobimalt to different binding sites on unfolded insulin. A similar explanation and the existence of such binding sites with different binding energies was shown previously for the nonionic detergent. Thus, the data also emphasize the importance of a protein partially-unfolded state which undergoes the process of fibrils formation; i.e., certain experimental conditions or the presence of additives may dramatically change not only kinetics but also the morphology of fibrillar aggregates.

2-Dimensional in vitro culture assessment of ovarian cancer cell line using cost effective silver nanoparticles from Macrotyloma uniflorum seed extracts.

Our research focused on generating AgNPs using Macrotyloma uniflorum (MU) seed extracts and studied their efficacy in combating tumor growth using the 2-Dimensional method for ovarian cancer cell line-PA-1. Characterization studies including a UV-visible spectrophotometer confirmed the surface plasmon resonance peak of 436 nm. Particle size determination data validated the nanoparticle diameter of 91.8 nm. Synthesized AgNPs possess a negative charge of -28.0 mV, which was confirmed through the zeta potential study. Structural characterization studies including XRD determined the crystal phase of AgNPs at four distant peaks at 2θ (38.17, 44.36, 64.52, and 77.46) and were assigned to 111, 200, 220, and 311 planes of the FCC. FTIR studies have confirmed the presence of O-H, N-H, C=O, ethers, C-Br, and C-I groups in AgNPs respectively. DPPH study has confirmed the presence of free radicles and we observed that at 500 μg/ml concentration, 76.08% of free radicles were formed which shows their efficiency. MTT assay shows the efficacy of MU-AgNPs in reducing the cell viability. At lower concentrations of MU-AgNP, 66% viability was observed and 9% of viability was observed at higher dose. ROS production (21%) was observed using MU-AgNPs with respect to 0.45% in controls, which affirms the capacity to induce DNA damage via apoptosis. Standard drug camptothecin generated 26% of ROS production which confirms higher potential of AgNPs in inducing DNA damage in tumor cells without causing lethality to the healthy cells. Further, the Fluorescence-activated cell sorting (FACS) study using a standard Caspase-3 marker confirms the generation of apoptotic bodies using two different concentrations of MU-AgNPs. At 40 μg, 64% of apoptotic cell death was observed, whereas, using 20 μg, 23% of apoptosis was recorded via fluorescent intensity. Propidium iodide-based Cell cycle study has shown a significant decrease in G0/G1 phase compared to control (88.8%), which further confirmed the apoptotic induction. Matrix metalloproteinases (MMP) studies using JC-1 dye, showed a significant increase in green fluorescence owing to lowered membrane potential, thus ensuring the breakdown of mitochondrial potential compared to untreated and standard drugs. With the obtained results, we are concluding that MU-AgNPs has a tremendous capacity to suppress the ovarian cancer cell proliferation in vitro by inducing DNA damage and apoptosis.

Zn2+-enhanced Ru (II) photoluminescence directed by double-clamp structural ligand for selective Zn2+ sensing and live-cell imaging

A novel water-soluble, visible-light-excitable, fluorescence enhancement sensor (λex = 450 nm), [Ru (PYPZA-tBuDO3A) (bipy)2]2+, has been synthesized and characterized, which efficiently recognize Zn2+ ions without interference from other biologically relevant metal ions, anions, amino acids, small biomolecules or pharmaceutical molecules in phosphate buffered saline (PBS, pH = 7.0). The detection has high sensitivity (detection limit = 5.3533 × 10-9 mol/L) for Zn2+ ions owning to the formation of 1:1 binding with a large binding constant (K = 2.1586 × 105 M-1). Density functional theory (DFT) and time-dependent density functional theory (TDDFT) calculations were performed to determine the mechanism for changes of photophysical properties illustrating fluorescence increase response to Zn2+ ions. The [Ru (PYPZA-tBuDO3A) (bipy)2]2+ also exhibits excellent membrane permeability and negligible cytotoxicity, and can be applied to sense intracellular Zn2+ in live human cells.