Methods and Applications in Fluorescence最新文献_第6页

Quasi-equilibrium state based quantification of biological macromolecules in single-molecule localization microscopy. 单分子定位显微镜中基于准平衡态的生物大分子定量。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-09-08 DOI: 10.1088/2050-6120/acf546

Xuecheng Chen, Yaqian Li, Xiaowei Li, Jielin Sun, Daniel M Czajkowsky, Zhifeng Shao

{"title":"Quasi-equilibrium state based quantification of biological macromolecules in single-molecule localization microscopy.","authors":"Xuecheng Chen, Yaqian Li, Xiaowei Li, Jielin Sun, Daniel M Czajkowsky, Zhifeng Shao","doi":"10.1088/2050-6120/acf546","DOIUrl":"https://doi.org/10.1088/2050-6120/acf546","url":null,"abstract":"The stoichiometry of molecular components within supramolecular biological complexes is often an important property to understand their biological functioning, particularly within their native environment. While there are well established methods to determine stoichiometryin vitro, it is presently challenging to precisely quantify this propertyin vivo,especially with single molecule resolution that is needed for the characterization stoichiometry heterogeneity. Previous work has shown that optical microscopy can provide some information to this end, but it can be challenging to obtain highly precise measurements at higher densities of fluorophores. Here we provide a simple approach using already established procedures in single-molecule localization microscopy (SMLM) to enable precise quantification of stoichiometry within individual complexes regardless of the density of fluorophores. We show that by focusing on the number of fluorophore detections accumulated during the quasi equilibrium-state of this process, this method yields a 50-fold improvement in precision over values obtained from images with higher densities of active fluorophores. Further, we show that our method yields more correct estimates of stoichiometry with nuclear pore complexes and is easily adaptable to quantify the DNA content with nanodomains of chromatin within individual chromosomes inside cells. Thus, we envision that this straightforward method may become a common approach by which SMLM can be routinely employed for the accurate quantification of subunit stoichiometry within individual complexes within cells.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10197268","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Rapid microwave synthesis of N and S dual-doped carbon quantum dots for natamycin determination based on fluorescence switch-off assay. 荧光关闭法快速微波合成N、S双掺杂碳量子点测定纳他霉素。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-08-25 DOI: 10.1088/2050-6120/acf119

Ali Abdel-Hakim, Fatallah Belal, Mohamed A Hammad, Mahmoud Hamed Elmaghrabey

{"title":"Rapid microwave synthesis of N and S dual-doped carbon quantum dots for natamycin determination based on fluorescence switch-off assay.","authors":"Ali Abdel-Hakim, Fatallah Belal, Mohamed A Hammad, Mahmoud Hamed Elmaghrabey","doi":"10.1088/2050-6120/acf119","DOIUrl":"https://doi.org/10.1088/2050-6120/acf119","url":null,"abstract":"Green, one-pot, quick, and easily synthesized nitrogen and sulfur co-doped carbon quantum dots (N,S-CDs) were obtained from cheap and readily available chemicals (sucrose, urea, and thiourea) using a microwave-assisted approach in about 4 min and utilized as a turn-off fluorescent sensor for estimation of natamycin (NAT). First, the effect of N and S doping on the microwave-synthesized CDs' quantum yield was carefully studied. CDs derived from sucrose alone failed to produce a high quantum yield; then, to increase the quantum yield, doping with heteroatoms was carried out using either urea or thiourea. A slight increase in quantum yield was observed upon using thiourea with sucrose, while an obvious enhancement of quantum yield was obtained when urea was used instead of thiourea. Surprisingly, using a combination of urea and thiourea together results in N,S-CDs with the highest quantum yield (53.5%), uniform and small particle size distribution, and extended stability. The fluorescent signal of N,S-CDs was quenched upon addition of NAT due to inner filter effect and static quenching in a manner that allowed for quantitative determination of NAT over a range of 0.5-10.0μg ml-1(LOD = 0.10μg ml-1). The N,S-CDs were applicable for determination of NAT in aqueous humor, eye drops, different environmental water samples, and bread with excellent performance. The selectivity study indicated excellent selectivity of the prepared N,S-CDs toward NAT with little interference from possibly interfering substances. In-silico toxicological evaluation of NAT was conducted to estimate its long-term toxicity and drug-drug interactions. Finally, the preparation of N,S-CDs, and analytical procedure compliance with the green chemistry principles were confirmed by two greenness assessment tools.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-08-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10080910","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

DWH24: a new antibody for fluorescence-based cell death analysis. DWH24:一种荧光细胞死亡分析的新抗体。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-08-23 DOI: 10.1088/2050-6120/aceed0

Anna Ryschich, Yan Dong, Michael Schäfer, Eduard Ryschich, Svetlana Karakhanova

{"title":"DWH24: a new antibody for fluorescence-based cell death analysis.","authors":"Anna Ryschich, Yan Dong, Michael Schäfer, Eduard Ryschich, Svetlana Karakhanova","doi":"10.1088/2050-6120/aceed0","DOIUrl":"https://doi.org/10.1088/2050-6120/aceed0","url":null,"abstract":"Antibodies have gained considerable importance in laboratory and clinical settings. Currently, antibodies are extensively employed for the diagnosis and treatment of several human diseases. Herein, using targeted and cell immunisation approaches, we developed and characterised an antibody clone, DWH24. We found that DWH24 is an IgMκtype antibody that enables excellent visualisation and quantification of dead cells using immunofluorescence, fluorescence microscopy, and flow cytometry. This property was proved by the spontaneous cell death of several tumour cell lines and stimulated T cells, as well as after chemo- and photodynamic therapy. Unlike conventional apoptosis and cell death markers, DWH24 binding occurred in a Ca2+- and protein-independent manner and enabled live imaging of cell death progress, as shown using time-lapse microscopy. The binding specificity of DWH24 was analysed using a human proteome microarray, which revealed a complex response profile with very high spot intensities against various proteins, such as tropomyosin variants and FAM131C. Accordingly, DWH24 can be employed as a suitable tool for the cost-effective and universal analysis of cell death using fluorescence imaging and flow cytometry.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-08-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10421226","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Spectrofluorometric determination of cinacalcet hydrochloride: greenness assessment and application to biological fluids andin-vitrodissolution testing. 荧光分光光度法测定盐酸西那卡塞的绿度评价及其在生物体液和体外溶出度试验中的应用。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-26 DOI: 10.1088/2050-6120/ace81b

Mona Abo Zaid, Nahed El Enany, Aziza E Mostafa, Ghada M Hadad, Fathalla Belal

{"title":"Spectrofluorometric determination of cinacalcet hydrochloride: greenness assessment and application to biological fluids andin-vitrodissolution testing.","authors":"Mona Abo Zaid, Nahed El Enany, Aziza E Mostafa, Ghada M Hadad, Fathalla Belal","doi":"10.1088/2050-6120/ace81b","DOIUrl":"https://doi.org/10.1088/2050-6120/ace81b","url":null,"abstract":"A facile, simple, green and sensitive spectrofluorometric method was developed for determination of the calcimimetic drug cinacalcet hydrochloride. It is used for the treatment of hyperparathyroidism. The drug showed high native fluorescence intensity at 320 nm after excitation at 280 nm. The method was linear over the range of 5.0-400.0 ng ml-1with excellent correlation (R2= 0.9999). Limit of detection (LOD) and limit of quantitation (LOQ) values were 1.19 and 3.62 ng ml-1, respectively. The percentage recovery was found to be 100.42% ± 1.39 (n=8). The proposed method was successfully applied for determination of cinacalcet in spiked human plasma samples with % recoveries of (87.23 to 109.69%). Two recent greenness metrics (GAPI and Analytical Eco-Scale) were chosen to prove the eco-friendly nature of the method. Furthermore, the proposed method was successfully applied to dissolution study of commercial cinacalcet tablets. The interference likely to be introduced by some commonly co-administrated drugs such as metoprolol and itraconazole was studied; the tolerance limits were calculated.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9928386","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Probing the potential toxicity of trimetazidine by characterizing its interaction with human serum albumin. 探讨曲美他嗪与人血清白蛋白相互作用的潜在毒性。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-19 DOI: 10.1088/2050-6120/ace513

Aya Barseem, Fathalla Belal, Mokhtar Mabrouk, Sherin Hammad, Hytham Ahmed

{"title":"Probing the potential toxicity of trimetazidine by characterizing its interaction with human serum albumin.","authors":"Aya Barseem, Fathalla Belal, Mokhtar Mabrouk, Sherin Hammad, Hytham Ahmed","doi":"10.1088/2050-6120/ace513","DOIUrl":"https://doi.org/10.1088/2050-6120/ace513","url":null,"abstract":"The small molecular drugs pharmacodynamics and pharmacokinetics could be affected by human serum albumin (HSA) transport, so we studied the interaction between HSA and the widely used anti-ischemic agent, trimetazidine (TMZ), using different approaches. As shown by synchronous fluorescence spectroscopy, the interaction affects the microenvironment confirmation around tyrosine residues. The site-competitive experiments showed that TMZ had an affinity toward subdomain III A (site II) of HSA. The enthalpy and entropy changes (ΔH and ΔS), which were 37.75 and 0.197 K J mol-1, respectively, showed that the predominant intermolecular interactions are hydrophobic forces. According to FTIR research, the interaction between HSA and TMZ caused polypeptide carbonyl-hydrogen bonds to rearrange. The HSA esterase enzyme activity was decreased with TMZ. Docking analysis confirmed the site-competitive experiments and thermodynamic results. This study demonstrated that TMZ interacted with HSA, and the structure and function of HSA were influenced by TMZ. This study could aid in understanding the pharmacokinetics of TMZ and provide basic data for safe use.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10219884","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Visualization of the composition of the urinary fluorescent metabolome. Why is it important to consider initial urine concentration? 尿荧光代谢组组成的可视化。为什么考虑初始尿浓度很重要?

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-19 DOI: 10.1088/2050-6120/ace512

Katarína Dubayová, Anna Birková, Martin Lešo, Jaroslava Žilková, Anton Karabinoš, Mária Mareková, Marek Stupak

引用次数: 0

Quasi-equilibrium state based quantification of biological macromolecules in single-molecule localization microscopy 单分子定位显微镜中基于准平衡态的生物大分子定量

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-18 DOI: 10.1101/2023.07.17.549270

Xuecheng Chen, Yaqian Li, Xiaowei Li, Jielin Sun, D. Czajkowsky, Zhifeng Shao

{"title":"Quasi-equilibrium state based quantification of biological macromolecules in single-molecule localization microscopy","authors":"Xuecheng Chen, Yaqian Li, Xiaowei Li, Jielin Sun, D. Czajkowsky, Zhifeng Shao","doi":"10.1101/2023.07.17.549270","DOIUrl":"https://doi.org/10.1101/2023.07.17.549270","url":null,"abstract":"The stoichiometry of molecular components within supramolecular biological complexes is often an important property to understand their biological functioning, particularly within their native environment. While there are well established methods to determine stoichiometry in vitro, it is presently challenging to precisely quantify this property in vivo, especially with single molecule resolution that is needed for the characterization stoichiometry heterogeneity. Previous work has shown that optical microscopy can provide some information to this end, but it can be challenging to obtain highly precise measurements at higher densities of fluorophores. Here we provide a simple approach using already established procedures in single-molecule localization microscopy (SMLM) to enable precise quantification of stoichiometry within individual complexes regardless of the density of fluorophores. We show that by focusing on the number of fluorophore detections accumulated during the quasi equilibrium-state of this process, this method yields a 50-fold improvement in precision over values obtained from images with higher densities of active fluorophores. Further, we show that our method yields more correct estimates of stoichiometry with nuclear pore complexes and is easily adaptable to quantify the DNA content with nanodomains of chromatin within individual chromosomes inside cells. Thus, we envision that this straightforward method may become a common approach by which SMLM can be routinely employed for the accurate quantification of subunit stoichiometry within individual complexes within cells.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-07-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"47048495","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Excited-state properties of 6-methoxyflavone in the presence of halide ions in aqueous media. 卤化物离子存在下6-甲氧基黄酮的激发态性质。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-07 DOI: 10.1088/2050-6120/ace152

Nisha Fatma, Sanjay Pant, Nupur Pandey, Mohan Singh Mehata

{"title":"Excited-state properties of 6-methoxyflavone in the presence of halide ions in aqueous media.","authors":"Nisha Fatma, Sanjay Pant, Nupur Pandey, Mohan Singh Mehata","doi":"10.1088/2050-6120/ace152","DOIUrl":"https://doi.org/10.1088/2050-6120/ace152","url":null,"abstract":"The present work investigated the influence of different halides on the excited state dynamics of 6-methoxyflavone (6MF) in an aqueous solution with steady-state and time-resolved techniques. On successive addition of I-and Br-ions, the fluorescence of 6MF quenched significantly, whereas the respective ions do not change the maximum fluorescence band. Fluorescence of 6MF was quenched 66% by I-ions and 34% by Br-ions. In a pure aqueous medium, both the H-bonded: CT and protonated species of 6MF participate in the quenching of fluorescence. The quenching process was categorized by Stern-Volmer (S-V) and Lehrer equations. Quenching parameters such as KSV, KSV-Land kqwere higher for I-ions than Br-ions. The decrease in fluorescence intensity and a reduction in fluorescence lifetime suggested the dynamic nature of quenching by I-ions following the electron transfer mechanism. Fluorescence quenching of 6MF has also been observed in the acidic medium in the presence of different halides. Thus, the study reveals that 6MF is responsive towards I-ions in a wide range of pH, specifically in a purely aqueous environment (pH∼7), hence important for sensing/detection applications.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-07-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10157473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Real-time correction of chromatic aberration in optical fluorescence microscopy. 光学荧光显微镜色差的实时校正。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-07-03 DOI: 10.1088/2050-6120/ace153

Ana Cayuela López, Pablo Conesa, Ana María Oña Blanco, José Antonio Gómez-Pedrero, Carlos Oscar S Sorzano

{"title":"Real-time correction of chromatic aberration in optical fluorescence microscopy.","authors":"Ana Cayuela López, Pablo Conesa, Ana María Oña Blanco, José Antonio Gómez-Pedrero, Carlos Oscar S Sorzano","doi":"10.1088/2050-6120/ace153","DOIUrl":"https://doi.org/10.1088/2050-6120/ace153","url":null,"abstract":"Multi-color fluorescence imaging is a powerful tool for studying the spatial relationships and interactions among sub-cellular structures in biological specimens. However, if improperly corrected, geometrical distortions caused by mechanical drift, refractive index mismatch, or chromatic aberration can lead to lower image resolution. In this paper, we present an extension of the image processing framework of Scipion by integrating a protocol called OFM Corrector, which corrects geometrical distortions in real-time using a B-spline-based elastic continuous registration technique. Our proposal provides a simple strategy to overcome chromatic aberration by digitally re-aligning color channels in multi-color fluorescence microscopy images, even in 3D or time. Our method relies on a geometrical calibration, which we do with fluorescent beads excited by different wavelengths of light and subsequently registered to get the elastic warp as a reference to correct chromatic shift. Our software is freely available with a user-friendly GUI and can be broadly used for various biological imaging problems. The paper presents a valuable tool for researchers working in light microscopy facilities.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"11 4","pages":""},"PeriodicalIF":3.2,"publicationDate":"2023-07-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9734822","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Two-photon lifetime-based photoconversion of EGFP for 3D-photostimulation in FLIM. 基于双光子寿命的EGFP在FLIM中用于3d光刺激的光转换。

IF 3.2 3区化学

Methods and Applications in Fluorescence Pub Date : 2023-06-12 DOI: 10.1088/2050-6120/acdb31

Dita Strachotová, Aleš Holoubek, Barbora Brodská, Petr Herman

引用次数: 0