Detection properties of indium-111 and IRDye800CW for intraoperative molecular imaging use across tissue phantom models.

Significance: Intraoperative molecular imaging (IMI) enables the detection and visualization of cancer tissue using targeted radioactive or fluorescent tracers. While IMI research has rapidly expanded, including the recent Food and Drug Administration approval of a targeted fluorophore, the limits of detection have not been well-defined. Aim: The ability of widely available handheld intraoperative tools (Neoprobe and SPY-PHI) to measure gamma decay and fluorescence intensity from IMI tracers was assessed while varying characteristics of both the signal source and the intervening tissue or gelatin phantoms. Approach: Gamma decay signal and fluorescence from tracer-bearing tumors (TBTs) and modifiable tumor-like inclusions (TLIs) were measured through increasing thicknesses of porcine tissue and gelatin in custom 3D-printed molds. TBTs buried beneath porcine tissue were used to simulate IMI-guided tumor resection. Results: Gamma decay from TBTs and TLIs was detected through significantly thicker tissue and gelatin than fluorescence, with at least 5% of the maximum signal observed through up to 5 and 0.5 cm, respectively, depending on the overlying tissue type or gelatin. Conclusions: We developed novel systems that can be fine-tuned to simulate variable tumor characteristics and tissue environments. These were used to evaluate the detection of fluorescent and gamma signals from IMI tracers and simulate IMI surgery.

Fluorescence and photoacoustic (FL/PA) dual-modal probe: Responsive to reactive oxygen species (ROS) for atherosclerotic plaque imaging.

Accurate and early detection of atherosclerosis (AS) is imperative for their effective treatment. However, fluorescence probes for efficient diagnosis of AS often encounter insufficient deep tissue penetration, which hinders the reliable assessment of plaque vulnerability. In this work, a reactive oxygen species (ROS) activated near-infrared (NIR) fluorescence and photoacoustic (FL/PA) dual model probe TPA-QO-B is developed by conjugating two chromophores (TPA-QI and O-OH) and ROS-specific group phenylboronic acid ester. The incorporation of ROS-specific group not only induces blue shift in absorbance, but also inhibits the ICT process of TPA-QO-OH, resulting an ignorable initial FL/PA signal. ROS triggers the convertion of TPA-QO-B to TPA-QO-OH, resulting in the concurrent amplification of FL/PA signal. The exceptional selectivity of TPA-QO-B towards ROS makes it effectively distinguish AS mice from the healthy. The NIR emission can achieve a tissue penetration imaging depth of 0.3 cm. Moreover, its PA775 signal possesses the capability to penetrate tissues up to a thickness of 0.8 cm, ensuring deep in vivo imaging of AS model mice in early stage. The ROS-triggered FL/PA dual signal amplification strategy improves the accuracy and addresses the deep tissue penetration problem simultaneously, providing a promising tool for in vivo tracking biomarkers in life science and preclinical applications.

Near-infrared fluorescent probe visual detection of Hg2+ and its application in biological system and ecological system.

Mercury ion (Hg2+), a heavy metal cation with greater toxicity, is widely present in the ecological environment and has become a serious threat to human health and environmental safety. Currently, developing a solution to simultaneously visualize and monitor Hg2+ in environmental samples, including water, soil, and plants, remains a great challenge. In this work, we created and synthesized a near-infrared fluorescent probe, BBN-Hg, and utilized Hg2+ to trigger the partial cleavage of the carbon sulfate ester in BBN-Hg as a sensing mechanism, and the fluorescence intensity of BBN-Hg was significantly enhanced at 650 nm, thus realizing the visualization of Hg2+ with good selectivity (detection limit, 53 nM). In live cells and zebrafish, the probe BBN-Hg enhances the red fluorescence signal in the presence of Hg2+, and successfully performs 3D imaging on zebrafish, making it a powerful tool for detecting Hg2+ in living systems. More importantly, with BBN-Hg, we are able to detect Hg2+ in actual water samples, soil and plant seedling roots. Furthermore, the probe was prepared as a test strip for on-site determination of Hg2+ with the assistance of a smartphone. Therefore, this study offers an easy-to-use and useful method for tracking Hg2+ levels in living organisms and their surroundings.

A multifunctional near-infrared fluorescent probe based on benzothiazole structure for fluoride-ion detection.

Fluoride ions (F-) are one of the essential trace elements for the human body and are widely existed in nature. In this study, we present a novel fluorescent probe (YF-SZ-F) designed and synthesized for the specific detection of F-. The probe exhibits high sensitivity, excellent selectivity, and low cytotoxicity, making it a promising tool for biomedical applications. Imaging experiments conducted on zebrafish and Arabidopsis roots demonstrate the probe's remarkable cellular permeability and biocompatibility, laying a solid foundation for its potential biomedical utility. Furthermore, the probe holds potential for practical applications in environmental monitoring and public health through its capability to detect fluoride ions in water samples and via mobile phone software. This multifaceted functionality underscores the broad applicability and significance of the fluorescent probe, not only in scientific research but also in real-world scenarios, contributing to the development of more convenient and precise methods for fluoride detection.

Developing an off-on fluorescence sensor based on red copper nanoclusters wrapped by sulfhydryl and polymer double ligands for sensitive detection of N-acetyl-L-cysteine.

N-acetyl-L-cysteine (NAC) as a class of thiols is commonly used in the treatment of lung diseases, detoxification and prevention of liver damage. In this paper, 4-mercaptobenzoic acid (4-MBA) coated and polyvinylpyrrolidone (PVP) attached copper nanoclusters (4-MBA@PVP-CuNCs) were successfully synthesized using a simple one-pot method with an absolute quantum yield of 10.98 %, and its synthetic conditions (like effects of single/double ligands and temperature) were studied intensively. Then Hg2+ could quench the fluorescence of the 4-MBA@PVP-CuNCs and its fluorescence was restored with the addition of NAC. Based on the above principles, an off-on switching system was established to detect NAC. That is, the 4-MBA@PVP-CuNCs-Hg probe was prepared by adding Hg2+ to switch off the fluorescence of the CuNCs by static quenching, and then NAC was added to switch on the fluorescence of the probe based on the chelation of NAC and Hg2+. Moreover, the effects of metal ion types and mercury ion doses for the probe construction were also further discussed. The method showed excellent linearity in the range of 0.05-1.25 µM and low detection limit of 16 nM. Meanwhile, good recoveries in real urine, tablets and pellets were observed, which proved the reliability of the method and provided a convenient, fast and sensitive method for NAC detection.

A near-infrared ratiometric fluorescent probe for the sensing and imaging of sulfur dioxide derivatives in living systems.

As a reactive sulfur species, sulfur dioxide (SO2) and its derivatives play crucial role in various physiological processes, which can maintain redox homeostasis at normal levels and lead to the occurrence of many diseases at abnormal levels. So, the development of a suitable fluorescent probe is a crucial step in advancing our understanding of the role of SO2 derivatives in living organisms. Herein, we developed a near-infrared fluorescent probe (SP) based on the ICT mechanism to monitor SO2 derivatives in living organisms in a ratiometric manner. The probe SP exhibited excellent selectivity, good sensitivity, fast response rate (within 50 s), and low detection limit (1.79 µM). In addition, the cell experiment results suggested that the SP has been successfully employed for the real-time monitoring of endogenous and exogenous SO2 derivatives with negligible cytotoxicity. Moreover, SP was effective in detecting SO2 derivatives in mice.

Synthesis and optical properties of tyramine-functionalized boron dipyrromethene dyes for cell bioimaging.

Tyramine signaling amplification (TSA) technology is generally applied in immunofluorescence, enzyme-linked immunoassays, in situ hybridization techniques, etc. Successful amplification of fluoresence signals cannot be achieved without excellent fluorescent dyes. BODIPY fluorophore is an ideal probe for cell fluorescence imaging, but pristine BODIPY cannot be direct used in the TSA system. In the paper, the new red-shifted tyramide-conjugated BODIPY (BDP-B/C/D) was synthesized via the Knoevenagel condensation reaction, which based on the tyramide-conjugated BODIPY (BDP-A). The synthesized dyes were combined with tyramine to obtain which could be used as a fluorescent substrate for enzymatic reaction of TSA. By using the selected substrate (BDP-C) in TSA, we found it to be more sensitive than the commercial dye 594 styramide for the detection of low-abundance antigen proteins.

Fluorescent probes based on N-CQDs: For direct detection of food additives STPP and Al3.

Sodium tripolyphosphate (STPP), as one of the many food additives, can cause gastrointestinal discomfort and a variety of adverse reactions when ingested by the human body, which is a great potential threat to human health. Therefore, it is necessary to develop a fast, sensitive and simple method to detect STPP in food. In this study, we synthesized a kind of nitrogen-doped carbon quantum dots (N-CQDs), and were surprised to find that the addition of STPP led to the gradual enhancement of the emission peaks of the N-CQDs, with a good linearity in the range of 0.067-1.96 µM and a low detection limit as low as 0.024 µM. Up to now, there is no report on the use of carbon quantum dots for the direct detection of STPP. Meanwhile, we found that the addition of Al3+ effectively bursts the fluorescence intensity of N-CQDs@STPP solution and has a good linear relationship in the range of 0.33-6.25 µM with a lower detection limit of 0.24 µM. To this end, we developed a fluorescent probe to detect STPP and Al3+. In addition, the probe was successfully applied to the detection of bread samples, which has great potential for practical application.

Sodium salicylate as a green fluorescent probe for ultrasensitive determination of vonoprazan fumarate via fluorescence switch off strategy; greenness assessment.

A green, simple and sensitive spectrofluorometric approach for determining vonoprazan fumarate in bulk and pharmaceutical dosage form by turning off the fluorescence of sodium salicylate is developed. The addition of vonoprazan fumarate reduced linearly the fluorescence intensity of 0.4 mM sodium salicylate at λem 408 nm and at λex 330 nm. The approach was found to be linear in the 50.0-3000.0 ng/mL range. The limits of detection and quantification were 10.97 and 33.23 ng/mL, respectively. The presented method proved its suitability in determination of vonoprazan fumarate in its pure and pharmaceutical dosage form. This method employs water as the exclusive solvent and utilizes safe reagents, evaluated using the Analytical Eco Scale, Green Analytical Procedure Index (GAPI), and carbon footprint. In contrast, previous methods relied on toxic reagents and required extended heating times, resulting in higher environmental impact. The novel method not only enhances analytical efficiency but also aligns with green chemistry principles, offering a sustainable solution for routine pharmaceutical analysis.

A dual-channel fluorescent probe targeting lysosomes for differential detection of Cys/Hcy and GSH: Applications in food, pharmaceutical analysis and bioimaging.

Thiols function as antioxidants in food, prolonging shelf life and enhancing flavor. Moreover, thiols are vital biomolecules involved in enzyme activity, cellular signal transduction, and protein folding among critical biological processes. In this paper, the fluorescent probe PYL-NBD was designed and synthesized, which utilized the fluorescent molecule pyrazoline, the lysosome-targeted morpholine moiety, and the sensing moiety NBD. Probe PYL-NBD was tailored for the recognition of biothiols through single-wavelength excitation, yielding distinct fluorescence emission signals: blue for Cys, Hcy, and GSH; green for Cys, Hcy. Probe PYL-NBD exhibited rapid reaction kinetics (<10 min), distinct fluorescence response signals, and low detection limits (15.7 nM for Cys, 14.4 nM for Hcy, and 12.6 nM for GSH). Probe PYL-NBD enabled quantitative determination of Cys content in food samples and L-cysteine capsules. Furthermore, probe PYL-NBD had been successfully applied for confocal imaging with dual-channel detection of biothiols in various biological specimens, including HeLa cells, zebrafish, tumor sections, and Arabidopsis thaliana.

Assessment of Viability of Listeria monocytogenes by Flow Cytometry.

In food industry, Listeria monocytogenes contamination can occur accidentally despite the quality control of raw materials and factory. Decontamination processes or inhibitory effects of ingredients/additives in food products are set up to ensure compliance with hygiene and microbiological criteria. These actions represent stresses for the pathogenic agent, causing fluctuations in its physiological states. Moreover, during these environmental stresses, Listeria monocytogenes can enter in a viable but nonculturable (VBNC) state which is not detected by plate counting but by flow cytometry. This technique coupled with cell staining by fluorescent dyes offers the possibility to assess different physiological states based on different cellular parameters: enzymatic activity, transmembrane integrity, membrane potential, and respiratory activity. In this chapter, we present a method to assess the viability of foodborne pathogens using a double-staining principle based on the assessment of membrane integrity and intracellular esterase activity.

Cysteine triggered cascade reaction forming coumarin: Visualization of cysteine fluctuation in alcoholic liver disease by a NIR fluorescent probe.

Alcoholic liver disease (ALD) is a chronic toxic liver injury caused by long-term heavy drinking. Due to the increasing incidence, ALD is becoming one of important medical tasks. Many studies have shown that the main mechanism of liver damage caused by large amounts of alcohol may be related to antioxidant stress. As an important antioxidant, cysteine (Cys) is involved in maintaining the normal redox balance and detoxifying metabolic function of the liver, which may be closely related to the pathogenesis of ALD. Therefore, it is necessary to develop a simple non-invasive method for rapid monitoring of Cys in liver. Thus, a near-infrared (NIR) fluorescent probe DCI-Ac-Cys which undergoes Cys triggered cascade reaction to form coumarin fluorophore is developed. Using the DCI-Ac-Cys, decreased Cys was observed in the liver of ALD mice. Importantly, different levels of Cys were monitored in the livers of ALD mice taking silybin and curcumin with the antioxidant effects, indicating the excellent therapeutic effect on ALD. This study provides the important references for the accurate diagnosis of ALD and the pharmacodynamic evaluation of silybin and curcumin in the treatment of ALD, and support new ideas for the pathogenesis of ALD.

The first ER-targeting flavone-based fluorescent probe for Cys: Applications in real-time tracking in an epilepsy model and food analysis.

Epilepsy is one of the most commonly-seen neurological disorders, and both endoplasmic reticulum stress (ERS) and oxidative stress (OS) have been demonstrated to be associated with epileptic seizures. As one of the three endogenous thiol-containing amino acids, cysteine (Cys) is recognized not only as an important biomarker of various biological processes but also widely used as a significant additive in the food industry. However, the exact role that Cys plays in ERS has not been well answered up to now. In this paper, we reported the first flavone-based fluorescent probe (namely BFC) with nice endoplasmic reticulum (ER)-targeting ability, which was capable of monitoring Cys in a fast response (3.0 min), large stokes shift (130 nm) and low detection limit (10.4 nM). The recognition mechanism of Cys could be attributed to the addition-cyclization reaction involving a Cys residue and an acrylate group, resulting in the release of the strong excited-state intramolecular proton transfer (ESIPT) emission molecule of benzoflavonol (BF). The low cytotoxicity and good biocompatibility of the probe BFC allowed for monitoring the fluctuation of endogenous Cys levels under both ERS and OS processes, as well as in zebrafish models of epilepsy. Quantitative determination of Cys with the probe BFC was also achieved in three different food samples. Additionally, a probe-immersed test strips integrated with a smartphone device was successfully constructed for on-site colorimetric detection of Cys. Undoubtedly, our work provided a valuable tool for tracking Cys levels in both an epilepsy model and real food samples.

AIE-based fluorescent probe designed with xanthone as a π-bridge for detecting of ClO- in pericarp and living cells.

The role of ClO- in the physiological functioning of organisms is significant. In this paper, the four fluorescent probes HONx (HON1, HON2, HON3 and HON4) were prepared based on oxyanthracene through the introduction of different substituents, and their photophysical properties were investigated, among which the AIE effect of HON1 was the most significant, and therefore the fluorescent "turn-off" ClO- probe HON1-CN was chosen to be prepared by constructing the ClO- recognition site hydrazone bond at HON1. The ClO- recognises the hydrazone group in the probe HON1-CN, and when the hydrazone bond is broken, the aldehyde group is released, generating HON1 with yellow fluorescence. The probe HON1-CN is highly selective and stable for the detection of ClO- with a detection limit of 0.48 µM and a more than 10-fold increase in fluorescence intensity when the fluorescence is 'switched on', and to a lesser extent, the probe is also very good for the detection of hypochlorite ClO- in the pericarp. Finally, HON1-CN has also been used to detect the presence of ClO- in HeLa cells and zebrafish.

Construction of eco-friendly dual carbon dots ratiometric fluorescence probe for highly selective and efficient sensing mercury ion.

In present work, blue carbon dots (b-CDs) were derived from ammonium citrate and guanidine hydrochloride, and red carbon dots (r-CDs) were stemmed from malonate, ethylenediamine and meso­tetra (4-carboxyphenyl) porphin based on facile hydrothermal method. Eco-friendly ratiometric fluorescence probe was innovatively constructed to effectively measure Hg2+ utilizing b-CDs and r-CDs. The developed probe displayed two typical emission peaks at 450 nm from b-CDs and 650 nm from r-CDs under the excitation at 360 nm. Mercury ion has strong quenching effect on the fluorescence intensity at 450 nm due to the electron transfer process and the fluorescence change at 450 nm was used as the response signal, whereas the fluorescence intensity at 650 nm kept unchangeable which resulted from the chemical inertness between Hg2+ and r-CDs, serving as the reference signal in the sensing system. Under optimal circumstances, this probe exhibited an excellent linearity between the fluorescence response values of ΔF450/F650 and Hg2+ concentrations over range of 0.01-10 µmol/L, and the limit of detection was down to 5.3 nmol/L. Furthermore, this probe was successfully employed for sensing Hg2+ in practical environmental water samples with satisfied recoveries of 98.5%-105.0%. The constructed ratiometric fluorescent probe provided a rapid, environmental-friendly, reliable, and efficient platform for measuring trace Hg2+ in environmental field.

Dual excitation channel ratiometric fluorescent probes for visual and fluorescent detection of anthrax spore biomarker and tetracycline hydrochloride.

Long-term and excessive use of tetracycline hydrochloride (TC) can lead to its accumulation in the environment, which can cause water contamination, bacterial resistance, and food safety problems. 2,6-Pyridine dicarboxylic acid (DPA) is a major biomarker of Bacillus anthracis spores, and its rapid and sensitive detection is of great significance for disease prevention and counter-terrorism. A bifunctional ratiometric fluorescent nanoprobe has been fabricated to detect DPA and TC. 3,5-dicarboxyphenylboronic acid (BOP) was intercalated into layered europium hydroxide (LEuH) by the ion-exchange method and exfoliated into nanosheets as a fluorescent nanoprobe (PNP). DPA and TC could significantly enhance the red fluorescence of Eu3+ through the antenna effect under different excitation wavelengths, while the fluorescence of BOP can be used as a reference based on the constant emission intensity, realizing ratiometric detection. A low limit of detection (LOD) for the target (DPA: 9.7 nM, TC: 21.9 nM) can be achieved. In addition, visual detection of DPA and TC was realized using color recognition software based on the obvious color changes. This is the first ratiometric fluorescent nanoprobe based on layered rare-earth hydroxide (LRH) for the detection of DPA and TC simultaneously, which opens new ideas in the design of multifunctional probes.

Highly selective and visual detection of vanillin based on fluorescence Cd-MOF sensor in milk powders.

Vanillin is a commonly used synthetic flavoring agent in daily life. However, excessive intake of vanillin may pose risks to human health. Therefore, there is an urgent need for rapid and sensitive detection methods for vanillin. In this study, we developed a fluorescent sensor based on Cd-MOF for the sensitive and selective recognition of vanillin. The presence of vanillin leads to significant fluorescence quenching of Cd-MOF due to competitive absorption and photoinduced electron transfer (PET). The limit of detection was determined to be 39.6 nM, which is the lowest-among the reported fluorescent probes. The sensor was successfully applied for the detection of vanillin in real samples such as powdered milk and milk, with a recovery rate ranging from 96.88 % to 104.83 %. Furthermore, by immobilizing the Cd-MOF probe into a polyvinyl alcohol (PVA) film, we achieved a portable and visual detection composite materials for vanillin.

Diagnostic value for methylation in cervical cancer based on a small-molecule fluorescent probe targeting DNMT1.

BACKGROUND: Value analysis of a small-molecule fluorescent probe for methylation detection in different cervical lesions. MATERIALS AND METHODS: (1) The grayscale values of distinct lesion tissues were remarkably distinct among the four groups (p < 0.05). The comparison of the grayscale value between the two groups showed that the CA group noticeably exceeded the LSIL and cervicitis groups, and the HSIL group was apparently higher than the LSIL and cervicitis groups (p < 0.05); (2) The mean grayscale values of the enrolled subjects were calculated with 55.21 as the midline, with >55.21 as positive and ≤55.21 as negative. RESULTS: The results showed that the positive rate of the cervicitis group was 0.00%, the LSIL group 67.74%, the HSIL group 83.33%, and the CA group 100.00%. The results among the four groups were notably distinct (p < 0.05); (3) The comparison among DAPI, probe, bright, and merged images of cervicitis, LSIL, HSIL, and CA indicated that different cervical lesions were with quite various stains. CONCLUSION: The grayscale value, positive rate, and stained picture of distinct cervical lesions were remarkably different. The small-molecule fluorescent probe has a good value in differentiating cervical lesions and can be considered for popularization and application.

High-throughput fluorescence lifetime imaging flow cytometry.

Flow cytometry is a vital tool in biomedical research and laboratory medicine. However, its accuracy is often compromised by undesired fluctuations in fluorescence intensity. While fluorescence lifetime imaging microscopy (FLIM) bypasses this challenge as fluorescence lifetime remains unaffected by such fluctuations, the full integration of FLIM into flow cytometry has yet to be demonstrated due to speed limitations. Here we overcome the speed limitations in FLIM, thereby enabling high-throughput FLIM flow cytometry at a high rate of over 10,000 cells per second. This is made possible by using dual intensity-modulated continuous-wave beam arrays with complementary modulation frequency pairs for fluorophore excitation and acquiring fluorescence lifetime images of rapidly flowing cells. Moreover, our FLIM system distinguishes subpopulations in male rat glioma and captures dynamic changes in the cell nucleus induced by an anti-cancer drug. FLIM flow cytometry significantly enhances cellular analysis capabilities, providing detailed insights into cellular functions, interactions, and environments.

Robust Double Emulsions for Multicolor Fluorescence-Activated Cell Sorting.

Cell-cell interactions are essential for the proper functioning of multicellular organisms. For example, T cells interact with antigen-presenting cells (APCs) through specific T-cell receptor (TCR)-antigen interactions during an immune response. Fluorescence-activated droplet sorting (FADS) is a high-throughput technique for efficiently screening cellular interaction events. Unfortunately, current droplet sorting instruments have significant limitations, most notably related to analytical throughput and complex operation. In contrast, commercial fluorescence-activated cell sorters offer superior speed, sensitivity, and multiplexing capabilities, although their use as droplet sorters is poorly defined and underutilized. Herein, we present a universally applicable and simple-to-implement workflow for generating double emulsions and performing multicolor cell sorting using a commercial FACS instrument. This workflow achieves a double emulsion detection rate exceeding 90%, enabling multicellular encapsulation and high-throughput immune cell activation sorting for the first time. We anticipate that the presented droplet sorting strategy will benefit cell biology laboratories by providing access to an advanced microfluidic toolbox with minimal effort and cost investment.