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1.
World J Gastrointest Surg ; 16(6): 1883-1893, 2024 Jun 27.
Article in English | MEDLINE | ID: mdl-38983339

ABSTRACT

BACKGROUND: Gastric cancer is a common malignant tumor of the digestive system worldwide, and its early diagnosis is crucial to improve the survival rate of patients. Indocyanine green fluorescence imaging (ICG-FI), as a new imaging technology, has shown potential application prospects in oncology surgery. The meta-analysis to study the application value of ICG-FI in the diagnosis of gastric cancer sentinel lymph node biopsy is helpful to comprehensively evaluate the clinical effect of this technology and provide more reliable guidance for clinical practice. AIM: To assess the diagnostic efficacy of optical imaging in conjunction with indocyanine green (ICG)-guided sentinel lymph node (SLN) biopsy for gastric cancer. METHODS: Electronic databases such as PubMed, Embase, Medline, Web of Science, and the Cochrane Library were searched for prospective diagnostic tests of optical imaging combined with ICG-guided SLN biopsy. Stata 12.0 software was used for analysis by combining the "bivariable mixed effect model" with the "midas" command. The true positive value, false positive value, false negative value, true negative value, and other information from the included literature were extracted. A literature quality assessment map was drawn to describe the overall quality of the included literature. A forest plot was used for heterogeneity analysis, and P < 0.01 was considered to indicate statistical significance. A funnel plot was used to assess publication bias, and P < 0.1 was considered to indicate statistical significance. The summary receiver operating characteristic (SROC) curve was used to calculate the area under the curve (AUC) to determine the diagnostic accuracy. If there was interstudy heterogeneity (I 2 > 50%), meta-regression analysis and subgroup analysis were performed. RESULTS: Optical imaging involves two methods: Near-infrared (NIR) imaging and fluorescence imaging. A combination of optical imaging and ICG-guided SLN biopsy was useful for diagnosis. The positive likelihood ratio was 30.39 (95%CI: 0.92-1.00), the sensitivity was 0.95 (95%CI: 0.82-0.99), and the specificity was 1.00 (95%CI: 0.92-1.00). The negative likelihood ratio was 0.05 (95%CI: 0.01-0.20), the diagnostic odds ratio was 225.54 (95%CI: 88.81-572.77), and the SROC AUC was 1.00 (95%CI: The crucial values were sensitivity = 0.95 (95%CI: 0.82-0.99) and specificity = 1.00 (95%CI: 0.92-1.00). The Deeks method revealed that the "diagnostic odds ratio" funnel plot of SLN biopsy for gastric cancer was significantly asymmetrical (P = 0.01), suggesting significant publication bias. Further meta-subgroup analysis revealed that, compared with fluorescence imaging, NIR imaging had greater sensitivity (0.98 vs 0.73). Compared with optical imaging immediately after ICG injection, optical imaging after 20 minutes obtained greater sensitivity (0.98 vs 0.70). Compared with that of patients with an average SLN detection number < 4, the sensitivity of patients with a SLN detection number ≥ 4 was greater (0.96 vs 0.68). Compared with hematoxylin-eosin (HE) staining, immunohistochemical (+ HE) staining showed greater sensitivity (0.99 vs 0.84). Compared with subserous injection of ICG, submucosal injection achieved greater sensitivity (0.98 vs 0.40). Compared with 5 g/L ICG, 0.5 and 0.05 g/L ICG had greater sensitivity (0.98 vs 0.83), and cT1 stage had greater sensitivity (0.96 vs 0.72) than cT2 to cT3 clinical stage. Compared with that of patients ≤ 26, the sensitivity of patients > 26 was greater (0.96 vs 0.65). Compared with the literature published before 2010, the sensitivity of the literature published after 2010 was greater (0.97 vs 0.81), and the differences were statistically significant (all P < 0.05). CONCLUSION: For the diagnosis of stomach cancer, optical imaging in conjunction with ICG-guided SLN biopsy is a therapeutically viable approach, especially for early gastric cancer. The concentration of ICG used in the SLN biopsy of gastric cancer may be too high. Moreover, NIR imaging is better than fluorescence imaging and may obtain higher sensitivity.

2.
World J Methodol ; 14(2): 92982, 2024 Jun 20.
Article in English | MEDLINE | ID: mdl-38983668

ABSTRACT

In accordance with the World Health Organization data, cancer remains at the forefront of fatal diseases. An upward trend in cancer incidence and mortality has been observed globally, emphasizing that efforts in developing detection and treatment methods should continue. The diagnostic path typically begins with learning the medical history of a patient; this is followed by basic blood tests and imaging tests to indicate where cancer may be located to schedule a needle biopsy. Prompt initiation of diagnosis is crucial since delayed cancer detection entails higher costs of treatment and hospitalization. Thus, there is a need for novel cancer detection methods such as liquid biopsy, elastography, synthetic biosensors, fluorescence imaging, and reflectance confocal microscopy. Conventional therapeutic methods, although still common in clinical practice, pose many limitations and are unsatisfactory. Nowadays, there is a dynamic advancement of clinical research and the development of more precise and effective methods such as oncolytic virotherapy, exosome-based therapy, nanotechnology, dendritic cells, chimeric antigen receptors, immune checkpoint inhibitors, natural product-based therapy, tumor-treating fields, and photodynamic therapy. The present paper compares available data on conventional and modern methods of cancer detection and therapy to facilitate an understanding of this rapidly advancing field and its future directions. As evidenced, modern methods are not without drawbacks; there is still a need to develop new detection strategies and therapeutic approaches to improve sensitivity, specificity, safety, and efficacy. Nevertheless, an appropriate route has been taken, as confirmed by the approval of some modern methods by the Food and Drug Administration.

3.
Sci Rep ; 14(1): 16233, 2024 Jul 14.
Article in English | MEDLINE | ID: mdl-39004658

ABSTRACT

Saline-sodic stress restricts the absorption of zinc by rice, consequently impacting the photosynthesis process of rice plants. In this experiment, Landrace 9 was selected as the test material and the potting method was employed to investigate the influence of ZnO nanoparticles (ZnO NPs) on zinc absorption and chlorophyll fluorescence in rice grown in saline-sodic land. The research findings demonstrate that the application of ZnO NPs proves to be more advantageous for the growth of rice in saline-sodic soil. Notably, the application of ZnO NPs significantly decreases the levels of Na+ and MDA in rice leaves in saline-sodic soil, while increasing the levels of K+ and Zn2+. Additionally, ZnO NPs enhances the content of chloroplast pigments, specific energy flux, quantum yield, and the performance of active PSII reaction center (PIABS) in rice leaves under saline-sodic stress. Furthermore, the relative variable fluorescence (WK and VJ) and quantum energy dissipation rate (φDo) of rice are also reduced. Therefore, the addition of ZnO NPs enhances the transfer of electrons and energy within the rice photosystem when subjected to saline-sodic stress. This promotes photosynthesis in rice plants growing in saline-sodic land, increasing their resistance to saline-sodic stress and ultimately facilitating their growth and development.


Subject(s)
Oryza , Photosynthesis , Plant Leaves , Soil , Zinc Oxide , Oryza/metabolism , Oryza/drug effects , Oryza/growth & development , Zinc Oxide/pharmacology , Photosynthesis/drug effects , Plant Leaves/metabolism , Plant Leaves/drug effects , Soil/chemistry , Chlorophyll/metabolism , Photosystem II Protein Complex/metabolism , Metal Nanoparticles/chemistry , Fluorescence , Salinity
4.
Methods Mol Biol ; 2839: 43-52, 2024.
Article in English | MEDLINE | ID: mdl-39008247

ABSTRACT

Characterizing the two- and three-dimensional distribution of trace metals in biological specimens is key to better understand their role in biological processes. Iron (Fe) is of particular interest in these trace metals due to its widespread role in maintaining cellular health and preventing disease. X-ray fluorescence microscopy (XFM) is emerging as the method of choice for investigators to interrogate the cellular and subcellular distribution of Fe. XFM utilizes the intrinsic X-ray fluorescence properties of each element to produce quantitative 2D and 3D distributions of trace metals within a sample. Herein, methods for sample preparation of cells and tissue for the determination of Fe distribution by XFM are described.


Subject(s)
Iron , Microscopy, Fluorescence , Iron/analysis , Iron/metabolism , Microscopy, Fluorescence/methods , Animals , Humans , Spectrometry, X-Ray Emission/methods , X-Rays
5.
Methods Mol Biol ; 2839: 77-97, 2024.
Article in English | MEDLINE | ID: mdl-39008249

ABSTRACT

Transmembrane transition metal transporter proteins are central gatekeepers in selectively controlling vectorial metal cargo uptake and extrusion across cellular membranes in all living organisms, thus playing key roles in essential and toxic metal homeostasis. Biochemical characterization of transporter-mediated translocation events and transport kinetics of redox-active metals, such as iron and copper, is challenged by the complexity in generating reconstituted systems in which vectorial metal transport can be studied in real time. We present fluorescence-based proteoliposome methods to monitor redox-active metal transmembrane translocation upon reconstitution of purified metal transporters in artificial lipid bilayers. By encapsulating turn-on/-off iron or copper-dependent sensors in the proteoliposome lumen and conducting real-time transport assays using small unilamellar vesicles (SUVs), in which selected purified Fe(II) and Cu(I) transmembrane importer and exporter proteins have been reconstituted, we provide a platform to monitor metal translocation events across lipid bilayers in real time. The strategy is modular and expandable toward the study of different transporter families featuring diverse metal substrate selectivity and promiscuity.


Subject(s)
Lipid Bilayers , Oxidation-Reduction , Proteolipids , Proteolipids/metabolism , Proteolipids/chemistry , Lipid Bilayers/metabolism , Lipid Bilayers/chemistry , Copper/metabolism , Copper/chemistry , Iron/metabolism , Metals/metabolism , Metals/chemistry , Biological Transport , Unilamellar Liposomes/metabolism , Unilamellar Liposomes/chemistry
6.
Virchows Arch ; 2024 Jul 09.
Article in English | MEDLINE | ID: mdl-38980338

ABSTRACT

Fluorescence confocal microscopy (FCM) is an optical technique that uses laser light sources of different wavelengths to generate real-time images of fresh, unfixed tissue specimens. FCM allows histological evaluation of fresh tissue samples without the associated cryo artifacts after frozen sectioning. The aim of this study was to prospectively evaluate pediatric tumor specimens and assess their suitability for fresh tumor sampling. In addition, we aimed to determine whether tumor cell isolation for stable cell culture is still feasible after FCM imaging. Pediatric tumor specimens were imaged using FCM. Tumor viability and suitability for tissue sampling were evaluated and compared with H&E staining after paraffin embedding. In addition, FCM-processed and non-FCM-processed tissue samples were sent for tumor cell isolation to evaluate possible effects after FCM processing. When comparing estimated tumor cell viability using FCM and H&E, we found good to excellent correlating estimates (intraclass correlation coefficient = 0.891, p < 0.001), as well as substantial agreement in whether the tissue appeared adequate for fresh tissue collection (κ = 0.762, p < 0.001). After FCM, seven out of eight samples yielded passable cell cultures, compared to eight out of eight for non-FCM processed samples. Our study suggests that the use of FCM in tumor sampling can increase the yield of suitable fresh tumor samples by identifying viable tumor areas and ensuring that sufficient tissue remains for diagnosis. Our study also provides first evidence that the isolation and growth of tumor cells in culture are not compromised by the FCM technique.

7.
J Biol Chem ; : 107528, 2024 Jul 01.
Article in English | MEDLINE | ID: mdl-38960038

ABSTRACT

Therapeutic inhibition of the viral protein Nef is an intriguing direction of antiretroviral drug discovery-it may revitalize immune mechanisms to target, and potentially clear, HIV-1-infected cells. Of the many cellular functions of Nef, the most conserved is the downregulation of surface CD4, which takes place through Nef hijacking the clathrin adaptor protein complex 2 (AP2)-dependent endocytosis. Our recent crystal structure has unraveled the molecular details of the CD4-Nef-AP2 interaction. Guided by the new structural knowledge, we have developed a fluorescence polarization-based assay for inhibitor screening against Nef's activity on CD4. In our assay, AP2 is included along with Nef to facilitate the proper formation of the CD4-binding pocket, and a fluorescently labeled CD4 cytoplasmic tail binds competently to the Nef-AP2 complex generating the desired polarization signal. The optimized assay has a good signal-to-noise ratio, excellent tolerance of DMSO and detergent, and the ability to detect competitive binding at the targeted Nef pocket, making it suitable for high-throughput screening.

8.
J Biol Chem ; : 107529, 2024 Jul 01.
Article in English | MEDLINE | ID: mdl-38960039

ABSTRACT

The multifunctional, HIV-1 accessory protein Nef enables infected cells to evade host immunity and thus plays a key role in viral pathogenesis. One prominent function of Nef is the downregulation of major histocompatibility complex class I (MHC-I), which disrupts antigen presentation and thereby allows the infected cells to evade immune surveillance by the cytotoxic T cells. Therapeutic inhibition of this Nef function is a promising direction of antiretroviral drug discovery as it may revitalize cytotoxic T cells to identify, and potentially clear, hidden HIV-1 infections. Guided by the crystal structure of the protein complex formed between Nef, MHC-I, and the hijacked clathrin adaptor protein complex 1 (AP1), we have developed a fluorescence polarization-based assay for inhibitor screening against Nef's activity on MHC-I. The optimized assay has a good signal-to-noise ratio, substantial tolerance of DMSO, and excellent ability to detect competitive inhibition, indicating that it is suitable for high-throughput screening.

9.
Luminescence ; 39(7): e4828, 2024 Jul.
Article in English | MEDLINE | ID: mdl-39004779

ABSTRACT

The antibiotic tetracycline can be efficiently used as medicine for the deterrence of bacterial infections in humans, animals, and plants. However, the unprecedented use of tetracycline is of great concern owing to its low biodegradability, extensive usage, and adverse impacts on the environment and water quality. In this study, a sensitive spectrofluorometric method was proposed for the direct determination of tetracycline, based on biocompatible fluorescent carbon dots (CDs). The synthesis of CDs was performed by adopting a green hydrothermal procedure from carrot juice without requiring surface passivation or outflowing any environmentally hazardous waste. X-ray diffraction analysis and transmission electron microscopy revealed amorphous spherical-shaped CDs that exhibited blue emission under blue illumination. The fabricated fluorescent probe directly detected tetracycline in the concentration range of 4.00 × 10-6 to 1.55 × 10-5 mol L-1 with an LOD of 1.33 × 10-6 mol L-1. The performance of the probe was assessed in a tap water sample, with recovery values between 80.70 and 103.60%. The method's greenness was evaluated using the Analytical Green metric approach (AGREE) and confirmed to be within the green range. The developed method is facile, rapid, cost-effective, and offers a wide linear range and satisfactory selectivity, making it potentially suitable for determining tetracycline in water applications.


Subject(s)
Anti-Bacterial Agents , Carbon , Daucus carota , Fluorescent Dyes , Fruit and Vegetable Juices , Quantum Dots , Spectrometry, Fluorescence , Tetracycline , Daucus carota/chemistry , Fluorescent Dyes/chemistry , Fluorescent Dyes/chemical synthesis , Quantum Dots/chemistry , Carbon/chemistry , Anti-Bacterial Agents/analysis , Tetracycline/analysis , Fruit and Vegetable Juices/analysis , Limit of Detection
10.
HardwareX ; 19: e00545, 2024 Sep.
Article in English | MEDLINE | ID: mdl-39006472

ABSTRACT

The development of a compact and affordable fluorescence microscope can be a formidable challenge for growing needs in on-site testing and detection of fluorescent labeled biological systems, especially for those who specialize in biology rather than in engineering. In response to such a situation, we present an open-source miniature fluorescence microscope using Raspberry Pi. Our fluorescence microscope, with dimensions of 19.2 × 13.6 × 8.2 cm3 (including the display, computer, light-blocking case, and other operational requirements), not only offers cost-effectiveness (costing less than $500) but is also highly customizable to meet specific application needs. The 12.3-megapixel Raspberry Pi HQ Camera captures high-resolution imagery, while the equipped wide-angle lens provides a field of view measuring 21 × 15 mm2. The integrated wireless LAN in the Raspberry Pi, along with software-controllable high-powered fluorescence LEDs, holds potential for a wide range of applications. This open-source fluorescence microscope offers biohybrid sensor developers a versatile tool to streamline unfamiliar mechanical design tasks and open new opportunities for on-site fluorescence detections.

11.
Cureus ; 16(6): e62212, 2024 Jun.
Article in English | MEDLINE | ID: mdl-39006698

ABSTRACT

Herein, we report a rare case of pleural epithelioid malignant mesothelioma with a prominent myxoid stroma. To date, detailed morphological or molecular pathological findings have not been reported for this type of tumor. Hence, we aimed to describe the cytological, histological, immuno-cytohistological, electron-microscopic, and molecular pathological findings using fluorescence in situ hybridization (FISH) in such a case. The patient was a male in his mid-sixties with a history of asbestos exposure and had originally visited the hospital with a persistent cough and fever. Chest radiography revealed left pleural effusion, and laboratory examination revealed a high titer for hyaluronic acid in the effusion. Additionally, computed tomography revealed diffuse multinodular or cystic lesions in the left parietal pleura, and pleural effusion cytology revealed large epithelioid cells with mild nuclear atypia, which were considered reactive mesothelial cells. Cytologically, Giemsa staining revealed that these cells harbored variously sized intracytoplasmic vacuoles that were Alcian-blue-positive, suggesting hyaluronan production. Biopsy revealed large epithelioid cells that loosely proliferated against a prominent myxoid background. These cells were immuno-positive for calretinin, Wilms' tumor 1, D2-40, vimentin, and cytokeratin AE1/AE3 but not for carcinoembryonic antigen, Ber-EP4, or desmin. BRCA 1 associated protein 1 immunostaining showed nuclear loss, and FISH showed homozygous deletion of cyclin-dependent kinase inhibitor 2A (p16) on chromosome 9p21. Based on these findings, the lesion was diagnosed as an epithelioid mesothelioma with a prominent myxoid stroma. Electron-microscopy demonstrated a dense microvillus pattern on the surface of the tumor cells, indicating a mesothelial cell origin, and variously sized vacuoles in the cytoplasm, confirming the presence of intracytoplasmic vacuoles demonstrated on cytology. The tumor tissues obtained during surgery harbored prominent myxoid stroma, which proved that the present tumor was consistent with this type of mesothelioma. After informed consent was obtained, the patient and family wished for total resection of the tumor and postoperative chemotherapy, and the patient eventually died eight months after surgery.

12.
Chemphyschem ; : e202400509, 2024 Jul 15.
Article in English | MEDLINE | ID: mdl-39007184

ABSTRACT

3-tolunitrile (3-TN) can from three different dimers, which differ in the relative orientation of the methyl groups. We determined the geometry changes of two of these conformers of 3-TN upon electronic excitation via a Franck-Condon fit of the vibronic intensities in the fluorescence emission spectra. Both aromatic rings expand upon electronic excitation, showing a delocalized one-photon excitation of the cluster. The conformer with the smaller COM distance shows an unusual order of the split components of the electronic origin. We attribute this changed order to the stronger CT contributions to the splitting and a partial breakdown of the point dipole approximation, made in the Frenkel type interaction.

13.
Water Sci Technol ; 90(1): 373-383, 2024 Jul.
Article in English | MEDLINE | ID: mdl-39007325

ABSTRACT

This study investigated the characteristics of dissolved organic matter (DOM) in two distinct water bodies, through the utilization of three-dimensional fluorescence spectroscopy coupled with self-organizing map (SOM) methodology. Specifically, this analysis concentrated on neurons 3, 14, and 17 within the SOM model, identifying notable differences in the DOM compositions of a coal subsidence water body (TX) and the MaChang Reservoir (MC). The humic substance content of DOM TX exceeded that of MC. The origin of DOM in TX was primarily linked to agricultural inputs and rainfall runoff, whereas the DOM in MC was associated with human activities, displaying distinctive autochthonous features and heightened biological activity. Principal component analysis revealed that humic substances dominated the DOM in TX, while the natural DOM in MC was primarily autochthonous. Furthermore, a multiple linear regression model (MLR) determined that external pollution was responsible for 99.11% of variation in the humification index (HIX) of water bodies.


Subject(s)
Humic Substances , Humic Substances/analysis , Organic Chemicals/analysis , Organic Chemicals/chemistry , Environmental Monitoring/methods , Spectrometry, Fluorescence/methods , Water Pollutants, Chemical/chemistry , Water Pollutants, Chemical/analysis , Principal Component Analysis
14.
Article in English | MEDLINE | ID: mdl-39007528

ABSTRACT

Conjugated polymer nanoparticles (CPNs or Pdots) have become increasingly popular fluorophores for multimodal applications that combine imaging with phototherapeutic effects. Reports of CPNs in photodynamic therapy applications typically focus on their ability to generate singlet oxygen. Alternatively, CPN excited states can interact with oxygen to form superoxide radical anion and a CPN-based hole polaron, both of which can have deleterious effects on fluorescence properties. Here, we demonstrate that CPNs prepared from the common conjugated polymer poly[(9,9-dioctylfluorenyl-2,7-diyl)-alt-co-(1,4-benzo-{2,1',3}-thiadiazole)] (PFBT, also known as F8BT) generate superoxide upon irradiation. We use the same CPNs to detect superoxide by doping them with a superoxide-responsive hydrocyanine dye developed by Murthy and co-workers. Superoxide induces off-to-on fluorescence switching by converting quenching hydrocyanine dyes to fluorescent cyanine dyes that act as fluorescence resonance energy transfer (FRET) acceptors for PFBT chromophores. Amplified FRET from the multichromophoric CPNs yields fluorescence signal intensities that are nearly 50 times greater than when the dye is excited directly or over 100 times greater when signal readout is from the CPN channel. The dye loading level governs the maximum amount of superoxide that induces a change in fluorescence properties and also influences the rate of superoxide generation by furnishing competitive excited state deactivation pathways. These results suggest that CPNs can be used to deliver superoxide in applications in which it is desirable and provide a caution for fluorescence-based CPN applications in which superoxide can damage fluorophores.

15.
J Fluoresc ; 2024 Jul 15.
Article in English | MEDLINE | ID: mdl-39007932

ABSTRACT

The binding properties between vitamin B12 (vitB12, cyanocobalamin) and fibrinogen (Fib) were investigated by UV-vis absorption and steady-state/three-dimentional (3D) fluorescence spectra techniques as well as molecular docking. The experimental results showed that the intrinsic fluorescence of Fib quenched by vitB12 with static mechanism to form a non-fluorescent complex. The positive signs of thermodynamic parameters, ΔH (92.18 kJ/mol) and ΔS (433.5 J/molK), indicated that the hydrophobic forces were dominant in the binding mode. The molecular docking data were found to be in agreement with these experimental results and were confirmed by three hydrophobic interactions between the Trp430, Try390 residues of Fib and the vitamin. 3D spectra showed that fibrinogen undergoes a conformation change when it interacts with vitB12. Based on non-radiative energy transfer theory, binding distance was calculated to be 3.94 nm between donor (tryptophan residues of Fib) and acceptor (vitB12). The limit of detection (LOD) of vitB12 was calculated as 2.08 µM in the presence of fibrinogen. The relative standard deviation (RSD) of method was 4.28% for determinations (n = 7) of a vitB12 solution with the concentration of 7.80 µM.

16.
Mikrochim Acta ; 191(8): 464, 2024 Jul 15.
Article in English | MEDLINE | ID: mdl-39007936

ABSTRACT

Rapid and high-sensitive Salmonella detection in milk is important for preventing foodborne disease eruption. To overcome the influence of the complex ingredients in milk on the sensitive detection of Salmonella, a dual-signal reporter red fluorescence nanosphere (RNs)-Pt was designed by combining RNs and Pt nanoparticles. After being equipped with antibodies, the immune RNs-Pt (IRNs-Pt) provide an ultra-strong fluorescence signal when excited by UV light. With the assistance of the H2O2/TMB system, a visible color change appeared that was attributed to the strong peroxidase-like catalytic activity derived from Pt nanoparticles. The IRNs-Pt in conjunction with immune magnetic beads can realize that Salmonella typhimurium (S. typhi) was captured, labeled, and separated effectively from untreated reduced-fat pure milk samples. Under the optimal experimental conditions, with the assay, as low as 50 CFU S. typhi can be converted to detectable fluorescence and absorbance signals within 2 h, suggesting the feasibility of practical application of the assay. Meanwhile, dual-signal modes of quantitative detection were realized. For fluorescence signal detection (emission at 615 nm), the linear correlation between signal intensity and the concentration of S. typhi was Y = 83C-3321 (R2 = 0.9941), ranging from 103 to 105 CFU/mL, while for colorimetric detection (absorbamce at 450 nm), the relationship between signal intensity and the concentration of S. typhi was Y = 2.9logC-10.2 (R2 = 0.9875), ranging from 5 × 103 to 105 CFU/mL. For suspect food contamination by foodborne pathogens, this dual-mode signal readout assay is promising for achieving the aim of convenient preliminary screening and accurate quantification simultaneously.


Subject(s)
Colorimetry , Milk , Salmonella typhimurium , Milk/microbiology , Milk/chemistry , Salmonella typhimurium/isolation & purification , Colorimetry/methods , Animals , Metal Nanoparticles/chemistry , Limit of Detection , Platinum/chemistry , Hydrogen Peroxide/chemistry , Fluorescence , Nanospheres/chemistry , Food Microbiology/methods , Food Contamination/analysis , Spectrometry, Fluorescence/methods
17.
Methods Mol Biol ; 2805: 203-212, 2024.
Article in English | MEDLINE | ID: mdl-39008184

ABSTRACT

Planarians are flatworms that have the remarkable ability to regenerate entirely new animals. This regenerative ability requires abundant adult stem cells called neoblasts, which are relatively small in size, sensitive to irradiation and the only proliferative cells in the animal. Despite the lack of cell surface markers, fluorescence-activated cell sorting (FACS) protocols have been developed to discriminate and isolate neoblasts, based on DNA content. Here, we describe a protocol that combines staining of far-red DNA dye Draq5, Calcein-AM and DAPI, along with a shortened processing time. This profiling strategy can be used to functionally characterize the neoblast population in pharmacologically-treated or gene knockdown animals. Highly purified neoblasts can be analyzed with downstream assays, such as in situ hybridization and RNA sequencing.


Subject(s)
Flow Cytometry , Planarians , Stem Cells , Animals , Planarians/cytology , Planarians/genetics , Flow Cytometry/methods , Stem Cells/cytology , Stem Cells/metabolism , Regeneration , Cell Separation/methods , Fluorescent Dyes/chemistry
18.
Biophys Chem ; 312: 107288, 2024 Jun 29.
Article in English | MEDLINE | ID: mdl-38991454

ABSTRACT

PADI4 is one of the human isoforms of a group of enzymes intervening in the conversion of arginine to citrulline. It is involved in the development of several types of tumors, as well as other immunological illnesses, such as psoriasis, multiple sclerosis, or rheumatoid arthritis. PADI4 auto-citrullinates in several regions of its sequence, namely in correspondence of residues Arg205, Arg212, Arg218, and Arg383. We wanted to study whether the citrullinated moiety affects the conformation of nearby regions and its binding to intact PADI4. We designed two series of synthetic peptides comprising either the wild-type or the relative citrullinated versions of such regions - i.e., a first series of peptides comprising the first three arginines, and a second series comprising Arg383. We studied their conformational properties in isolation by using fluorescence, far-ultraviolet (UV) circular dichroism (CD), and 2D1H NMR. Furthermore, we characterized the binding of the wild-type and citrullinated peptides in the two series to the intact PADI4, by using isothermal titration calorimetry (ITC), fluorescence, and biolayer interferometry (BLI), as well as by molecular docking simulations. We observed that citrullination did not alter the local conformational propensities of the isolated peptides. Nevertheless, for all the peptides in the two series, citrullination slowed down the kinetic koff rates of the binding reaction to PADI4, probably due to differences in electrostatic effects compared to the presence of arginine. The affinities of PADI4 for unmodified peptides were slightly larger than those of the corresponding citrullinated ones in the two series, but they were all within the same range, indicating that there were no relevant variations in the thermodynamics of binding due to sequence effects. These results highlight details of the self-citrullination of PADI4 and, more generally, of possible auto-catalytic mechanisms taking place in vivo for other citrullinating enzymes or, alternatively, in proteins undergoing citrullination passively.

19.
Spectrochim Acta A Mol Biomol Spectrosc ; 322: 124782, 2024 Jul 05.
Article in English | MEDLINE | ID: mdl-38991616

ABSTRACT

In this study, an innovative ratiometric fluorescence and smartphone-assisted visual sensing platform based on blue-yellow dual-emission carbon dots (BY-CDs) was constructed for the first time to determine brilliant blue. The BY-CDs was synthesized via a facile one-step hydrothermal process involving propyl gallate and o-phenylenediamine. The synthesized BY-CDs exhibit favorable water solubility and exceptional fluorescence stability. Under excitation at 370 nm, BY-CDs show two distinguishable fluorescence emission bands (458 and 558 nm). Upon addition of brilliant blue, the fluorescence intensity at 558 nm exhibited a significant quenching effect attributed to fluorescence resonance energy transfer (FRET), while the fluorescence intensity at 458 nm was basically unchanged. The prepared BY-CDs can effectively serve as a ratiometric nanosensor for determining brilliant blue with the ratio of fluorescence intensities at 458 and 558 nm (F458/F558) as response signal. In addition, the developed ratiometric fluorescence sensor exhibits a noticeable alteration in color from yellow to green under UV light with a wavelength of 365 nm upon addition of varying concentrations of brilliant blue, which provides the possibility of visual detection of brilliant blue by a smartphone application. Finally, the BY-CDs based dual-mode sensing platform successfully detected brilliant blue in actual food samples and achieved a desirable recovery rate. This study highlights the merits of fast, convenient, economical, real-time, visual, high accuracy, excellent precision, good selectivity and high sensitivity for brilliant blue detection, and paves new paths for the monitoring of brilliant blue in real samples.

20.
Spectrochim Acta A Mol Biomol Spectrosc ; 322: 124797, 2024 Jul 08.
Article in English | MEDLINE | ID: mdl-38991618

ABSTRACT

Discrimination of segmented Baijiu contributes to stabilizing the quality of products, improving revenue-generating effects. A fluorescence sensor array is constructed based on four fluorescence characteristic peaks of terbium@lanthanum metal-organic framework (Tb@La-MOF). Its fluorescence signal is specifically quenched, when Tb@La-MOF encounters acetaldehyde. Acetaldehyde may inhibit the absorption of energy by the organic ligands in MOF, or/and hydrogen bonding with -COOH on the organic ligand, resulting in energy transfer to Tb(Ⅲ). According to this, the quantitative detection of acetaldehyde is completed with a range of 10-300 µM and the detection limit of 5.5 µM. At the same time, it has been successfully applied to the discrimination of segmented Baijiu. Fifteen segmented from three wine cellars are 100 % discriminated with the combined processing of sensor arrays and analytical methods. Accuracy, simplicity, and low-cost are highlights of this fluorescence sensor array, which has considerable potential for application in detection, production, and food field.

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