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1.
Food Chem ; 398: 133879, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-35987002

RESUMO

Accurate and real-time reporting the freshness and quality of meat has far-reaching implications for safeguarding food safety and public health. Owing to the significant indicator role of H2S in meat spoilage, we here designed a ''dual-key-and-lock'' kind of H+-powered H2S-responsive ratiometric fluorescent nanoprobe BODIPY/Cy7Cl@HyNPs for precise and real-time evaluating the meat freshness. After incubation with spoilt meat, BODIPY/Cy7Cl@HyNPs was simultaneously initiated by the abnormal H+ and H2S acted as "dual-keys". As a result, the fluorescence emission of BODIPY/Cy7Cl@HyNPs at âˆ¼ 688 nm was enhanced, and the other emission at âˆ¼ 818 nm was quenched, thereby yielding a typical H+-powered ratiometric fluorescence response toward H2S in meat. In addition, we also successfully exploited BODIPY/Cy7Cl@HyNPs to achieve the real-time monitoring of the meat freshness during storage by measuring the changes of fluorescence signals and solution color.


Assuntos
Corantes , Carne , Fluorescência , Corantes Fluorescentes
2.
Methods Mol Biol ; 2564: 47-52, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36107336

RESUMO

Many fluorescent proteins are available nowadays suitable for in vivo imaging experiments. Each fluorescent protein has unique biophysical properties, such as emission and excitation spectra, quantum yield, oligomerization state, pH sensitivity, fluorescence lifetime, and stability within the cellular environment. Even a small variation in fluorescent protein properties might result in significant differences in the experimental outcomes. This chapter discusses the aspects that need to be considered while selecting the fluorescent proteins for in vivo imaging experiment.


Assuntos
Diagnóstico por Imagem , Proteínas , Biofísica , Corantes , Diagnóstico por Imagem/métodos , Fluorescência
3.
Methods Mol Biol ; 2564: 53-74, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36107337

RESUMO

Fluorescent proteins are standard tools for addressing biological questions in a cell biology laboratory. The genetic tagging of protein of interest with fluorescent proteins opens the opportunity to follow them in vivo and to understand their interactions and dynamics. In addition, the latest advances in optical microscopy image acquisition and processing allow us to study many cellular processes in vivo. Techniques such as fluorescence lifetime microscopy and hyperspectral imaging provide valuable tools for understanding fluorescent protein interactions and their photophysics. Finally, fluorescence fluctuation analysis opens the possibility to address questions of molecular diffusion, protein-protein interactions, and oligomerization, among others, yielding quantitative information on the subject of study. This chapter will cover some of the more important advances in cutting-edge technologies and methods that, combined with fluorescent proteins, open new frontiers for biological studies.


Assuntos
Corantes , Proteínas , Fenômenos Fisiológicos Celulares , Microscopia de Fluorescência/métodos
4.
Methods Mol Biol ; 2564: 99-119, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36107339

RESUMO

Fluorescent proteins have revolutionized cell biology and cell imaging through their use as genetically encoded tags. Structural biology has been pivotal in understanding how their unique fluorescent properties manifest through the formation of the chromophore and how the spectral properties are tuned through interaction networks. This knowledge has in turn led to the construction of novel variants with new and improved properties. Here we describe the process by which fluorescent protein structures are determined, starting from recombinant protein production to structure determination by molecular replacement. We also describe how to incorporate and determine the structures of proteins containing non-natural amino acids. Recent advances in protein engineering have led to reprogramming of the genetic code to allow incorporation of new chemistry at designed residue positions, with fluorescent proteins being at the forefront of structural studies in this area. The impact of such new chemistry on protein structure is still limited; the accumulation of more protein structures will undoubtedly improve our understanding and ability to engineer proteins with new chemical functionality.


Assuntos
Aminoácidos , Código Genético , Aminoácidos/química , Corantes , Cristalização , Engenharia de Proteínas/métodos , Proteínas Recombinantes/genética
5.
Methods Mol Biol ; 2564: 213-222, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36107344

RESUMO

Fluorescent proteins within fluorescent fusions have been reported to affect cellular growth fitness via altering native protein function and intracellular localization. Here we report in detail a procedure to analyze the growth characteristics of yeast cells expressing such fusions in comparison to unmodified parental strain. This approach can serve as an initial step in fluorescent protein characterization in vivo.


Assuntos
Proteínas de Saccharomyces cerevisiae , Saccharomycetales , Corantes/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomycetales/metabolismo
6.
Eur J Histochem ; 66(1)2022 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-35212500

RESUMO

The spread technique proposed by Miller and Beatty in 1969 allowed for the first time the visualization at transmission electron microscopy of nucleic acids and chromatin in an isolated and distended conformation. The final step of staining the spread chromatin is of critical importance because it can strongly influence the interpretation of the results. We evaluated different staining techniques and the most part of them provided a good result. Specifically, well contrasted micrographs were obtained when staining with H3PW12O40 (PTA), as originally proposed by Miller and Beatty, and with two alternatives proposed here: uranyl acetate or terbium citrate staining. Quite good contrast of the spread DNA could be achieved also by using Osmium Ammine; while no or few contrast of nucleic acids was observed by staining with KMnO4 and H3PMo12O40 (PMA) respectively.


Assuntos
Ácidos Nucleicos , Cromatina , Corantes , Microscopia Eletrônica , Coloração e Rotulagem
7.
Sci Rep ; 12(1): 15213, 2022 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-36075955

RESUMO

The color of yarn dyed woven fabrics comes from a series of different colored yarns mixed in complex ways and various proportions. Thus, predicting the color mixing effect or formulating the recipe is a difficult task which should consider the interaction between colored yarns and structure variations. Color mixing recipes, which are also called color prediction models, for woven fabrics have generally been derived through the two-dimensional modeling of woven structures. In this study, the three-dimensional geometrical and colorimetrical modelings of single-, double-, and three-layered woven fabrics in a wide range of colors were conducted to obtain two types of optimal spectral reflectance-based color prediction models. Through model evaluations, the obtained three-dimensional color prediction models were proved to have much higher predictive accuracy, especially in terms of lightness and chroma predictions, than that of the two-dimensional color prediction models previously developed.


Assuntos
Corantes , Têxteis
8.
Molecules ; 27(17)2022 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-36080461

RESUMO

A number of new 2-methyl- and 2-arylvinyl-3-nitropyridines were synthesized and their reactions with thiols were studied. It was found that 3-NO2 tends to be selectively substituted under the action of sulfur nucleophiles in the presence of another nucleofuge in position 5. Correlations between the substitution pattern and regioselectivity as well as photophysical properties were established. Some synthesized compounds possessed a large Stokes shift.


Assuntos
Corantes , Compostos de Sulfidrila
9.
Cir. Esp. (Ed. impr.) ; 100(9): 534-554, sept. 2022. tab
Artigo em Espanhol | IBECS | ID: ibc-ADZ-897

RESUMO

El verde de indocianina es una tinción fluorescente visible con luz cercana al infrarrojo. Es útil para la identificación de las estructuras anatómicas (tracto biliar, uréteres, paratiroides, conducto torácico), la vascularización de tejidos (en anastomosis en cirugía colorrectal, esofágica, gástrica, bariátrica, para plastias y colgajos en cirugía de pared abdominal, hepática, en hernias estranguladas en la isquemia intestinal), para la identificación de tumores (hígado, páncreas, suprarrenal, implantes en la carcinomatosis peritoneal, tumores retroperitoneales y linfomas) y para la identificación del ganglio centinela y del mapeo linfático de tumores malignos (cáncer de estómago, mama, colon, recto, esófago y piel). Las evidencias son muy alentadoras, aunque se necesita la estandarización de su uso y más estudios prospectivos y aleatorizados con mayor número de pacientes para obtener conclusiones definitivas sobre su uso. El objetivo de esta revisión de la literatura es proveer una guía para el uso de la fluorescencia con verde de indocianina en procedimientos de cirugía general (AU)


Indocyanine Green is a fluorescent substance visible in near-infrared light. It is useful for the identification of anatomical structures (biliary tract, ureters, parathyroid, thoracic duct), the tissues vascularization (anastomosis in colorectal, esophageal, gastric, bariatric surgery, for plasties and flaps in abdominal wall surgery, liver resection, in strangulated hernias and in intestinal ischemia), for tumor identification (liver, pancreas, adrenal glands, implants of peritoneal carcinomatosis, retroperitoneal tumors and lymphomas) and sentinel node identification and lymphatic mapping in malignant tumors (stomach, breast, colon, rectum, esophagus and skin cancer). The evidence is very encouraging, although standardization of its use and randomized studies with higher number of patients are required to obtain definitive conclusions on its use in general surgery. The aim of this literature review is to provide a guide for the use of ICG fluorescence in general surgery procedures (AU)


Assuntos
Humanos , Verde de Indocianina/administração & dosagem , Corantes/administração & dosagem , Procedimentos Cirúrgicos Operatórios/métodos
10.
J Hazard Mater ; 439: 129683, 2022 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-36104909

RESUMO

The water pollution caused by heavy metals and dyes emitted by industries has become a worldwide problem. These pollutants are difficult to be biodegraded. Even at low concentrations, they are toxic and at last threaten human health. Herein, while using Tetrahymena thermophila, a single-celled ciliate protozoa, to enrich and remove the heavy metal Cd2+ from water, CdS nanoparticle-Tetrahymena thermophila hybrid system (CdS-T. thermophila) for dye pollution remediation under light irradiation was developed. The conditions of Cd2+ enrichment and removal by T. thermophila, construction of efficient CdS-T. thermophila, and decolorization of Congo red using CdS-T. thermophila were investigated. In the presence of cysteine ethyl ester, the removal rate of Cd2+ by T. thermophila was 94% at low Cd2+ concentration of 1 mg L-1. The adsorption capacity of T. thermophila to Cd2+ reached 43 mg g-1 at Cd2+ concentration of 80 mg L-1. Using 0.1 g L-1 constructed CdS-T. thermophila, the decolorization rate of 50 mg L-1 Congo red solution reached 95% in 60 min under light irradiation. This study provides a new insight to effective removing Cd2+ from water by T. thermophila to construct the CdS-T. thermophila and using it to remediate dye pollution in the environment.


Assuntos
Metais Pesados , Tetrahymena thermophila , Adsorção , Cádmio/metabolismo , Corantes/metabolismo , Vermelho Congo/metabolismo , Humanos , Metais Pesados/metabolismo , Água
11.
Int J Mol Sci ; 23(17)2022 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-36077048

RESUMO

In view of the water pollution issues caused by pathogenic microorganisms and harmful organic contaminants, nontoxic, environmentally friendly, and efficient antimicrobial agents are urgently required. Herein, a nickel-based Keggin polyoxomolybdate [Ni(L)(HL)]2H[PMo12O40] 4H2O (1, HL = 2-acetylpyrazine thiosemicarbazone) was prepared via a facile hydrothermal method and successfully characterized. Compound 1 exhibited high stability in a wide range of pH values from 4 to 10. 1 demonstrated significant antibacterial activity, with minimum inhibitory concentration (MIC) values in the range of 0.0019-0.2400 µg/mL against four types of bacteria, including Staphylococcus aureus (S. aureus), Bacillus subtilis (B. subtilis), Escherichia coli (E. coli), and Agrobacterium tumefaciens (A. tumefaciens). Further time-kill studies indicated that 1 killed almost all (99.9%) of E. coli and S. aureus. Meanwhile, the possible antibacterial mechanism was explored, and the results indicate that the antibacterial properties of 1 originate from the synergistic effect between [Ni(L)(HL)]+ and [PMo12O40]3-. In addition, 1 presented effective adsorption of basic fuchsin (BF) dyes. The kinetic data fitted a pseudo-second-order kinetic model well, and the maximum adsorption efficiency for the BF dyes (29.81 mg/g) was determined by the data fit of the Freundlich isotherm model. The results show that BF adsorption was dominated by both chemical adsorption and multilayer adsorption. This work provides evidence that 1 has potential to effectively remove dyes and pathogenic bacteria from wastewater.


Assuntos
Níquel , Purificação da Água , Adsorção , Antibacterianos/química , Antibacterianos/farmacologia , Corantes/farmacologia , Escherichia coli , Níquel/química , Corantes de Rosanilina/farmacologia , Staphylococcus aureus
12.
Int J Mol Sci ; 23(17)2022 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-36077071

RESUMO

Concerns about the health of the planet have grown dramatically, and the dyeing sector of the textile industry is one of the most polluting of all industries. Nanoclays can clean dyeing wastewater using their adsorption capacities. In this study, as a new finding, it was possible to analyze and quantify the amount of metal ions substituted by anionic dyes when adsorbed, and to determine the optimal amount of nanoclay to be used to adsorb all the dye. The tests demonstrated the specific amount of nanoclay that must be used and how to optimize the subsequent processes of separation and processing of the nanoclay. Hydrotalcite was used as the adsorbent material. Direct dyes were used in this research. X-ray diffraction (XRD) patterns allowed the shape recovery of the hydrotalcite to be checked and confirmed the adsorption of the dyes. An FTIR analysis was used to check the presence of characteristic groups of the dyes in the resulting hybrids. The thermogravimetric (TGA) tests corroborated the dye adsorption and the thermal fastness improvement. Total solar reflectance (TSR) showed increased radiation protection for UV-VIS-NIR. Through the work carried out, it has been possible to establish the maximum adsorption point of hydrotalcite.


Assuntos
Hidróxido de Magnésio , Poluentes Químicos da Água , Adsorção , Hidróxido de Alumínio , Corantes , Têxteis
13.
Int J Mol Sci ; 23(17)2022 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-36077196

RESUMO

Herein, a novel laccase gene, Melac13220, was amplified from Methylobacterium extorquens and successfully expressed in Escherichia coli with a molecular weight of approximately 50 kDa. The purified Melac13220 had no absorption peak at 610 nm and remained silent within electron paramagnetic resonance spectra, suggesting that Melac13220 belongs to the non-blue laccase group. Both inductively coupled plasma spectroscopy/optical emission spectrometry (ICP-OES) and isothermal titration calorimetry (ITC) indicated that one molecule of Melac13220 can interact with two iron ions. Furthermore, the optimal temperature of Melac13220 was 65 °C. It also showed a high thermolability, and its half-life at 65 °C was 80 min. Melac13220 showed a very good acid environment tolerance; its optimal pH was 1.5. Cu2+ and Co2+ can slightly increase enzyme activity, whereas Fe2+ could increase Melac13220's activity five-fold. Differential scanning calorimetry (DSC) indicated that Fe2+ could also stabilize Melac13220. Unlike most laccases, Melac13220 can efficiently decolorize Congo Red and Indigo Carmine dyes even in the absence of a redox mediator. Thus, the non-blue laccase from Methylobacterium extorquens shows potential application value and may be valuable for environmental protection, especially in the degradation of dyes at low pH.


Assuntos
Lacase , Methylobacterium extorquens , Corantes/química , Escherichia coli/metabolismo , Concentração de Íons de Hidrogênio , Índigo Carmim , Lacase/metabolismo , Methylobacterium extorquens/metabolismo , Temperatura
14.
Org Lett ; 24(36): 6686-6691, 2022 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-36053069

RESUMO

The synthesis of aniline derivatives, common building blocks in many pharmaceuticals, agrochemicals, dyes or polymers, has been limited to reactions based on benzene-toluene-xylene derivatives (BTX) due to their ample availability. Despite the large number of existing methodologies, the synthesis of chiral 4-(sec-alkyl)anilines has not been accomplished so far. In this work, a tandem strategy based on the generation of a reactive aza-p-quinone methide (aza-p-QM) intermediate followed by Cu(I)-catalyzed addition of Grignard reagents has been developed.


Assuntos
Benzeno , Xilenos , Agroquímicos , Compostos de Anilina , Catálise , Corantes , Indicadores e Reagentes , Indolquinonas , Preparações Farmacêuticas , Polímeros , Tolueno
15.
BMJ Open ; 12(9): e061829, 2022 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-36100304

RESUMO

INTRODUCTION: Nowadays, two predominant methods for detecting sentinel lymph nodes (SLNs) in cervical cancer are in use. The most conventional method is a combination of a radiotracer, technetium-99m (99mTc) and blue dye. More recently, another method for SLN mapping using indocyanine green (ICG) is becoming widely accepted. ICG is a fluorescent dye, visualised intraoperatively with near-infrared (NIR) fluorescence imaging, providing real-time visual navigation. The presumed advantages of ICG over 99mTc, that is, being cheaper, non-radioactive and logistically more attractive, are only valuable if its detection rate proves to be at least non-inferior. Before omitting the well-functioning and evidence-based combined approach of 99mTc and blue dye, we aim to provide prospective evidence on the non-inferiority of ICG with NIR fluorescence imaging. METHODS AND ANALYSIS: We initiated a prospective non-inferiority study with a paired comparison of both SLN methods in a single sample of 101 patients with International Federation of Gynecology and Obstetrics (FIGO) stage IA-IB2 or IIA1 cervical cancer receiving primary surgical treatment. All patients undergo SLN mapping with ICG and NIR fluorescence imaging in adjunct to mapping with 99mTc (including single photon emission computed tomography with X-ray computed tomography (SPECT/CT)) and blue dye. Surgeons start SLN detection with ICG while being blinded for the preoperative outcome of SPECT/CT to avoid biased detection with ICG. Primary endpoint of this study is bilateral SLN detection rate of both methods (ie, detection of at least one SLN in each hemipelvis). Since we compare strategies for SLN mapping that are already applied in current daily practice for different types of cancer, no additional risks or burdens are expected from these study procedures. ETHICS AND DISSEMINATION: The current study is approved by the Medical Ethics Research Committee Utrecht (reference number 21-014). Findings arising from this study will be disseminated in peer-reviewed journals, academic conferences and through patient organisations. TRIAL REGISTRATION NUMBER: NL9011 and EudraCT 2020-005134-15.


Assuntos
Linfadenopatia , Linfonodo Sentinela , Neoplasias do Colo do Útero , Corantes , Feminino , Humanos , Verde de Indocianina , Estudos Prospectivos , Linfonodo Sentinela/diagnóstico por imagem , Linfonodo Sentinela/patologia , Linfonodo Sentinela/cirurgia , Biópsia de Linfonodo Sentinela/métodos , Tecnécio , Neoplasias do Colo do Útero/diagnóstico por imagem , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/cirurgia
16.
Sci Rep ; 12(1): 15499, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-36109538

RESUMO

The jeopardy of the synthetic dyes effluents on human health and the environment has swiftly aggravated, threatening human survival. Hence, sustained studies have figured out the most acceptable way to eliminate this poisonous contaminant. Thereby, our investigation aimed to fabricate montmorillonite/magnetic NiFe2O4@amine-functionalized chitosan (MMT-mAmCs) composite as a promising green adsorbent to remove the cationic methylene blue (MB) and crystal violet (CV) dyes from the wastewater in neutral conditions. Interestingly, MMT-mAmCs composite carries high negative charges at a wide pH range from 4 to 11 as clarified from zeta potential measurements, asserting its suitability to adsorb the cationic contaminants. In addition, the experimental study confirmed that the optimum pH to adsorb both MB and CV was pH 7, inferring the ability of MMT-mAmCs to adsorb both cationic dyes in simple process conditions. Furthermore, the ferromagnetic behavior of the MMT-mAmCs composite is additional merit to our adsorbent that provides facile, fast, and flawless separation. Notably, the as-fabricated composite revealed an auspicious adsorbability towards the adsorptive removal of MB and CV, since the maximum adsorption capacity of MB and CV were 137 and 118 mg/g, respectively. Moreover, the isotherm and kinetic investigatins depicted that the adsorption of both cationic dyes fitted Langmuir and Pseudo 2nd order models, respectively. Besides, the advanced adsorbent preserved satisfactory adsorption characteristics with maximal removal efficacy exceeding 87% after reuse for ten consecutive cycles. More importantly, MMT-mAmCs efficiently adsorbed MB and CV from real agricultural water, Nile river water and wastewater samples at the neutral pH medium, reflecting its potentiality to be a superb reusable candidate for adsorptive removal cationic pollutants from their aquatic media.


Assuntos
Quitosana , Poluentes Ambientais , Purificação da Água , Aminas , Bentonita/química , Cátions , Corantes/química , Violeta Genciana , Humanos , Azul de Metileno/química , Compostos Organometálicos , Águas Residuárias/química , Água
17.
Methods Mol Biol ; 2543: 1-11, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36087254

RESUMO

This chapter describes a simple, nondestructive, annexin V apoptosis detection method that can be employed in real time over a 48-h test exposure. The real-time functionality allows for temporal resolution of apoptotic and cell death responses during the test exposure and obviates the need for onerous sample preparation and time course protocols associated with other annexin V methods. Further, this technique is eminently accessible to a wide range of laboratories because it does not require flow cytometry or other cytometric methods. It was developed for use with a variety of microplate well densities and with standard multimodal plate readers. The central feature of this assay is that it continuously reports the residency status of phosphatidylserine (PS) on the exofacial surface of a cell as it is translocated from the inner membrane leaflet during the apoptotic process. This homogenous, no-wash assay is made possible by two optimized and distinct annexin V fusion proteins which contain complementing NanoBiT™ luciferase enzyme subunits, a time-released luciferase substrate, and a fluorescent membrane integrity reagent. During an apoptotic event, the luminescent signal arising from an assay well is proportional to the number of cells with PS exposure, and fluorescence intensity correlates with the degree of cell death (secondary necrosis). Conversely, untreated cells contribute negligible luminescent or fluorescent signals throughout the time course. The data collected from these assay measures provide for both standard potency determinations and kinetic characterization of dose- and agent-dependent apoptotic responses, from early through late phases.


Assuntos
Apoptose , Fosfatidilserinas , Anexina A5/metabolismo , Apoptose/fisiologia , Corantes , Citometria de Fluxo/métodos , Humanos , Necrose , Fosfatidilserinas/metabolismo
18.
J Am Podiatr Med Assoc ; 112(2)2022 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-36115037

RESUMO

Despite advancements in the treatment of diabetic patients with "at-risk" limbs, minor and major amputations remain commonplace. The diabetic population is especially prone to surgical complications from lower extremity amputation because of comorbidities such as renal disease, hypertension, hyperlipidemia, microvascular and macrovascular disease, and peripheral neuropathy. Complication occurrence may result in increases in hospital stay duration, unplanned readmission rate, mortality rate, number of operations, and incidence of infection. Skin flap necrosis and wound healing delay secondary to inadequate perfusion of soft tissues continues to result in significant morbidity, mortality, and cost to individuals and the health-care system. Intraoperative indocyanine green fluorescent angiography for the assessment of tissue perfusion may be used to assess tissue perfusion in this patient population to minimize complications associated with amputations. This technology provides real-time functional assessment of the macrovascular and microvascular systems in addition to arterial and venous flow to and from the flap soft tissues. This case study explores the use of indocyanine green fluorescent angiography for the treatment of a diabetic patient with a large dorsal and plantar soft-tissue deficit and need for transmetatarsal amputation with nontraditional rotational flap coverage. The authors theorize that the use of indocyanine green may decrease postoperative complications and cost to the health-care system through fewer readmissions and fewer procedures.


Assuntos
Verde de Indocianina , Retalhos Cirúrgicos , Amputação/métodos , Angiografia/métodos , Corantes , Humanos , Complicações Pós-Operatórias/etiologia , Retalhos Cirúrgicos/irrigação sanguínea
19.
J Virol Methods ; 309: 114610, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36064127

RESUMO

Inactivation of human respiratory viruses in air and on surfaces is important to control their spread. Exposure to germicidal ultraviolet (UV-C) light damages viral nucleic acid rendering them non-infectious. Most of the recent viral inactivation studies have not considered potential artifacts caused by interactions between UV-C light and culture media used to suspend and deposit virus on surfaces. We show that the reactive oxygen and nitrogen species (ROS and RNS) form when commonly used virus culture media is exposed to 265 nm irradiation from light emitting diodes (LEDs) at UV-C doses (4 or 40 mJ/cm2) commonly considered to achieve multiple log-inactivation of virus. Surface viral inactivation values were enhanced from 0.49 to 2.92 log10 of viruses in DMEM, EMEM or EMEM-F as compared to absence of culture media (only suspended in Tris-buffer). The mechanisms responsible for the enhanced surface inactivate is hypothesized to involve photo-activation of vitamins and dyes present in the culture media, deposited with the virus on surfaces to be disinfected, which produce ROS and RNS. Given the rapidly growing research and commercial markets for UV-C disinfecting devices, there is a need to establish surface disinfecting protocols that avoid viral inactivation enhancement artifacts associated with selection and use of common cell culture media in the presence of UV-C light. This study addresses this weak link in the literature and highlights that inadequate selection of virus suspension media may cause a bias (i.e., over-estimation) for the UV-C dosages required for virus inactivation on surfaces.


Assuntos
Ácidos Nucleicos , Vírus , Viés , Técnicas de Cultura de Células , Corantes , Meios de Cultura , Desinfecção/métodos , Humanos , Nitrogênio , Oxigênio , Espécies Reativas de Oxigênio , Raios Ultravioleta , Inativação de Vírus/efeitos da radiação , Vitaminas
20.
Nat Commun ; 13(1): 5363, 2022 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-36097007

RESUMO

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 ß-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.


Assuntos
Diagnóstico por Imagem , Sistemas do Segundo Mensageiro , Animais , Corantes , Proteínas de Fluorescência Verde/genética , Células HEK293 , Humanos , Camundongos
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