Methods and Applications in Fluorescence最新文献

{"title":"Fluorescent cysteine probe based on a signal amplification unit, a catalyzed hairpin assembly reaction and Förster resonance energy transfer","authors":"Sirirat Ouiganon, Chongdee Thammakhet-Buranachai, P. Thavarungkul, P. Kanatharana, C. Buranachai","doi":"10.1088/2050-6120/ac6664","DOIUrl":"https://doi.org/10.1088/2050-6120/ac6664","url":null,"abstract":"This work developed a sensitive DNA-based fluorescent probe comprising a cysteine binding unit and a signal amplification unit based on a catalyzed hairpin assembly (CHA) reaction. The cysteine binding unit comprises a homodimer of single-stranded DNA (ssDNA) rich in cytosine and held together by silver ions. In the presence of cysteine, the homodimer is disintegrated because of cysteine-silver binding that liberates the ssDNA, which drives the CHA reaction in the signal amplification unit. Förster resonance energy transfer (FRET) was used to report the generation of the amplified double-stranded DNA (dsDNA) product. Under the optimal conditions, the probe provided a good linearity (100–1200 nM), a good detection limit (47.8 ± 2.7 nM) and quantification limit (159.3 ± 5.3 nM), and a good sensitivity (1.900 ± 0.045 μM−1). The probe was then used to detect cysteine in nine real food supplement samples. All results provided good recoveries that are acceptable by the AOAC, indicating that it has potential for practical applications.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-04-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46475986","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}

{"title":"Phasor-based multi-harmonic unmixing for in-vivo hyperspectral imaging","authors":"A. Vallmitjana, Paola Lepanto, F. Irigoín, Leonel Malacrida","doi":"10.1101/2022.03.31.486485","DOIUrl":"https://doi.org/10.1101/2022.03.31.486485","url":null,"abstract":"Hyperspectral imaging (HSI) is a paramount technique in biomedical science, however, unmixing and quantification of each spectral component is a challenging task. Traditional unmixing relies on algorithms that need spectroscopic parameters from the fluorescent species in the sample. The phasor-based multi-harmonic unmixing method requires only the empirical measurement of the pure species to compute the pixel-wise photon fraction of every spectral component. Using simulations, we demonstrate the feasibility of the approach for up to 5 components and explore the use of adding a 6th unknown component representing autofluorescence. The simulations show that the method can be successfully used in typical confocal imaging experiments (with pixel photon counts between 101 and 103). As a proof of concept, we tested the method in living cells, using 5 common commercial dyes for organelle labeling and we easily and accurately separate them. Finally, we challenged the method by introducing a solvatochromic probe, 6-Dodecanoyl-N,N-dimethyl-2-naphthylamine (LAURDAN), intended to measure membrane dynamics on specific subcellular membrane-bound organelles by taking advantage of the linear combination between the organelle probes and LAURDAN. We succeeded in monitoring the membrane order in the Golgi apparatus, Mitochondria, and plasma membrane in the same in-vivo cell and quantitatively comparing them. The phasor-based multi-harmonic unmixing method can help expand the outreach of HSI and democratize its use by the community for it does not require specialized knowledge.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42961900","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}

{"title":"Analysis of constituents present in smokeless tobacco (<i>Nicotiana tabacum)</i>using spectroscopic techniques.","authors":"Pratima Mishra, Rohit Kumar, Abhishek Dwivedi, Awadhesh Kumar Rai","doi":"10.1088/2050-6120/ac5e11","DOIUrl":"10.1088/2050-6120/ac5e11","url":null,"abstract":"<p><p>Laser-Induced Breakdown Spectroscopy (LIBS) is an analytical technique used to identify and quantify the elements present in any type of material present in any phase (solid, liquid, gas, and aerosol). In the present work, our objective is to find the presence of toxic and other elements in chewing tobacco (<i><b>Nicotiana tabacum</b></i>) using LIBS. Spectral signatures of elements like C, Fe, Si, Mg, Mn, Ca, Ti, Na, H, N, K, O, along with some toxic elements Al, Sr, Li, Cu, Sb, and Cr are observed in the LIBS spectra of these tobacco samples. The spectral intensity ratio is measured for quantitative analysis of elements present in the samples. Further, Atomic Absorption Spectroscopy is used for determining absolute concentration in these samples. A relation between the AAS result and the relative intensity of spectral lines measured in the LIBS is obtained using regression analysis. The multivariate technique, Principal Component Analysis (PCA), discriminates all the samples based on their toxicity and other constituents. Molecular study (Photoacoustic spectroscopy (PAS), UV-Visible (UV-vis), and FT-IR) of tobacco samples were performed to analyze the molecules present in the tobacco samples.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"60498616","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}

{"title":"Lifetime based axial contrast enable simple 3D-STED imaging","authors":"Yuanqing Ma, Alex Macmillan, Ying Yang, K. Gaus","doi":"10.1088/2050-6120/ac5e10","DOIUrl":"https://doi.org/10.1088/2050-6120/ac5e10","url":null,"abstract":"Stimulated Emission Depletion (STED) microscopy increase spatial image resolution by laterally sharpening the illumination profile of the confocal microscope. However, it remains compromised in axial resolution. To improve axial STED resolution, constructive interference of the STED depletion beam must be formed surrounding the focal plane to turn off the fluorophores beyond the focal plane. For isotropic 3D-STED resolution, this axial STED interference pattern must be overlayed with the doughnut STED beam at nanometer accuracy. Such optical configurations can be challenging in alignment. In this current work, we introduced a straightforward lifetime based axial contrast in STED microscope by imaging the samples on an ITO coated glass coverslip. The STED laser generates surface plasmon resonance on the ITO surface that enhanced the metal induced energy transfer MIET effect on the ITO surface. The enhanced MIET effect established a lifetime gradient with ∼20% dynamic range that extend for mor than 400 nm from the ITO surface. The axial contrast based on the lifetime gradient was directly used for 3D-STED imaging of tubulin fibers inside COS-7 cells, where the vertical displacement of single tubulin fiber was revealed. Lifetime gating could be applied to further improve lateral spatial resolution. Considering that most common implementation of STED microscopes uses pulsed lasers and timing electronics, there is no optical modification of the microscope is required in the current 3D-STED approach.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-03-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42305795","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}

{"title":"Microwave-assisted synthesis and upconversion luminescence of NaYF4:Yb, Gd, Er and NaYF4:Yb, Gd, Tm nanorods.","authors":"Shivanand H Nannuri,&nbsp;Simranjit Singh,&nbsp;Superb K Misra,&nbsp;Santhosh C,&nbsp;Sajan D George","doi":"10.1088/2050-6120/ac58e6","DOIUrl":"https://doi.org/10.1088/2050-6120/ac58e6","url":null,"abstract":"<p><p>Anisotropic rare earth ion (RE³⁺) doped fluoride upconversion particles are emerging as potential candidate in diverse areas, ranging from biomedical imaging to photonics. Here, we develop a facile strategy to synthesize NaYF₄: Yb, Gd, Er, and NaYF₄: Yb, Gd, Tm upconversion nanorods via microwave synthesis route by controlling the synthesis time and compared the optical properties similar nanorods prepared via solvothermal technique. With the increase in synthesis time, the phase of the particle found to change from mixed phase to purely hexagonal and morphology of the particles change mixed phase of spherical and rod-shaped particles to completely nanorods for a synthesis time of 60 min. Further, the intrinsically hydrophobic particles changed to hydrophilic by removal of oleic capping via acid treatment and the amine functionalized silica coating. The upconversion luminescence as well as laser power dependent emission properties of the surface modified particles elucidate that surface modification route influence the upconversion luminescence as well as solvent dependent emission properties. Moreover, the laser power dependent studies elucidate that the upconversion process in a multi-photon process.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 2","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39822539","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}

{"title":"Enhanced fluorescence from semiconductor quantum dot-labelled cells excited at 280 nm.","authors":"Mollie McFarlane,&nbsp;Nicholas Hall,&nbsp;Gail McConnell","doi":"10.1088/2050-6120/ac5878","DOIUrl":"https://doi.org/10.1088/2050-6120/ac5878","url":null,"abstract":"<p><p>Semiconductor quantum dots (QDs) have significant advantages over more traditional fluorophores used in fluorescence microscopy including reduced photobleaching, long-term photostability and high quantum yields, but due to limitations in light sources and optics, are often excited far from their optimum excitation wavelengths in the deep-UV. Here, we present a quantitative comparison of the excitation of semiconductor QDs at a wavelength of 280 nm, compared to the longer wavelength of 365 nm, within a cellular environment. We report increased fluorescence intensity and enhanced image quality when using 280 nm excitation compared to 365 nm excitation for cell imaging across multiple datasets, with a highest average fluorescence intensity increase of 3.59-fold. We also find no significant photobleaching of QDs associated with 280 nm excitation and find that on average, ∼80% of cells can tolerate exposure to high-intensity 280 nm irradiation over a 6-hour period.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 2","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39648379","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}

{"title":"Study of synthesis temperature effect on β-NaGdF4: Yb3+, Er3+ upconversion luminescence efficiency and decay time using maximum entropy method","authors":"D. Pominova, I. Romanishkin, V. Proydakova, S. Kuznetsov, P. Grachev, A. Ryabova, N. Tabachkova, P. Fedorov, V. Loschenov","doi":"10.1088/2050-6120/ac5bdc","DOIUrl":"https://doi.org/10.1088/2050-6120/ac5bdc","url":null,"abstract":"Upconversion materials have several advantages for many applications due to their great potential in converting infrared light to visible. For practical use, it is necessary to achieve high intensity of UC luminescence, so the studies of the optimal synthesis parameters for upconversion nanoparticles are still going on. In the present work, we analyzed the synthesis temperature effect on the efficiency and luminescence decay of β-NaGd0.78Yb0.20Er0.02F4 (15–25 nm) upconversion nanoparticles with hexagonal crystal structure synthesized by anhydrous solvothermal technique. The synthesis temperature was varied in the 290 °C–320 °C range. The synthesis temperature was shown to have a significant influence on the upconversion luminescence efficiency and decay time. The coherent scattering domain linearly depended on the synthesis temperature and was in the range 13.1–22.3 nm, while the efficiency of the upconversion luminescence increases exponentially from 0.02 to 0.10% under 1 W cm−2 excitation. For a fundamental analysis of the reasons for the upconversion luminescence intensity dependence on the synthesis temperature, it was proposed to use the maximum entropy method for luminescence decay kinetics processing. This method does not require a preliminary setting of the number of exponents and, due to this, makes it possible to estimate additional components in the luminescence decay kinetics, which are attributed to different populations of rare-earth ions in different conditions. Two components in the green luminescence and one component in the red luminescence decay kinetics were revealed for nanoparticles prepared at 290 °C–300 °C. An intense short and a weak long component in green luminescence decay kinetics could be associated with two different populations of ions in the surface quenching layer and the crystal core volume. With an increase in the synthesis temperature, the second component disappears, and the decay time increases due to an increase in the number of ions in the crystal core volume and a more uniform distribution of dopants.","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":" ","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"44254974","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}

{"title":"-808 nm-activated Ca²⁺doped up-conversion nanoparticles that release no inducing liver cancer cell (HepG2) apoptosis.","authors":"Xinmeng Fa,&nbsp;Shaowei Lin,&nbsp;Jianghua Yang,&nbsp;Chong Shen,&nbsp;Yuanli Liu,&nbsp;Yongyang Gong,&nbsp;Aimiao Qin,&nbsp;Jun Ou,&nbsp;Ute Resch-Genger","doi":"10.1088/2050-6120/ac5524","DOIUrl":"https://doi.org/10.1088/2050-6120/ac5524","url":null,"abstract":"<p><p>A near-infrared (NIR) light-triggered release method for nitric oxide (NO) was developed utilizing core/shell NaYF₄: Tm/Yb/Ca@NaGdF₄: Nd/Yb up-conversion nanoparticles (UCNPs) bearing a mesoporous silica (mSiO₂) shell loaded with the NO donor S-nitroso-N-acetyl-DL-penicillamine (SNAP). To avoid overheating in biological samples, Nd³⁺was chosen as a sensitizer, Yb³⁺ions as the bridging sensitizer, and Tm³⁺ions as UV-emissive activator while co-doping with Ca²⁺was done to enhance the luminescence of the activator Tm³⁺. NO release from SNAP was triggered by an NIR-UV up-conversion process, initiated by 808 nm light absorbed by the Nd³⁺ions. NO release was confirmed by the Griess method. Under 808 nm irradiation, the viability of the liver cancer cell line HepG2 significantly decreased with increasing UCNPs@mSiO₂-SNAP concentration. For a UCNPs@mSiO₂-SNAP concentration of 200<i>μ</i>g ml^-1, the cell survival probability was 47%. These results demonstrate that UCNPs@mSiO₂-SNAP can induce the release of apoptosis-inducing NO by NIR irradiation.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 2","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-02-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39787958","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}

{"title":"Introducing the multi-dimensional spectral phasors: a tool for the analysis of fluorescence excitation-emission matrices.","authors":"L B P Socas,&nbsp;E E Ambroggio","doi":"10.1088/2050-6120/ac5389","DOIUrl":"https://doi.org/10.1088/2050-6120/ac5389","url":null,"abstract":"<p><p>The use of phasors to analyze fluorescence data was first introduced for time-resolved studies for a simpler mathematical analysis of the fluorescence-decay curves. Recently, this approach was extended to steady-state experiments with the introduction of the spectral phasors (SP), derived from the Fourier transform of the fluorescence emission spectrum. In this work, we revise key mathematical aspects that lead to an interpretation of SP as the characteristic function of a probability distribution. This formalism allows us to introduce a new tool, called multi-dimensional spectral phasor (MdSP) that seize, not only the information from the emission spectrum, but from the full excitation-emission matrix (EEM). In addition, we developed a homemade open-source Java software to facilitate the MdSP data processing. Due to this mathematical conceptualization, we settled a mechanism for the use of MdSP as a tool to tackle spectral signal unmixing problems in a more accurate way than SP. As a proof of principle, with the use of MdSP we approach two important biophysical questions: protein conformational changes and protein-ligand interactions. Specifically, we experimentally measure the EEM changes upon denaturation of human serum albumin (HSA) or during its association with the fluorescence dye 1,8-anilinonaphtalene sulphate (ANS) detected via tryptophan-ANS Förster Resonance Energy Transfer (FRET). In this sense, MdSP allows us to obtain information of the system in a simpler and finer way than the traditional SP. Specifically, understanding a protein's EEM as a molecular fingerprint opens new doors for the use of MdSP as a tool to analyze and comprehend protein conformational changes and interactions.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 2","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-02-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39903698","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}

{"title":"A new near-infrared fluorescent probe for sensing extreme acidity and bioimaging in lysosome.","authors":"Qiuchen Liu,&nbsp;Chang Liu,&nbsp;Songtao Cai,&nbsp;Song He,&nbsp;Liancheng Zhao,&nbsp;Xianshun Zeng,&nbsp;Jin Zhou,&nbsp;Jin Gong","doi":"10.1088/2050-6120/ac4e73","DOIUrl":"https://doi.org/10.1088/2050-6120/ac4e73","url":null,"abstract":"<p><p>Since the intracellular pH plays an important role in the physiological and pathological processes, however, the probes that can be used for monitoring pH fluctuation under extreme acidic conditions are currently rare, so it is necessary to construct fluorescent probes for sensing pH less than 4. In this work, we developed a new near-infrared (NIR) fluorescent probe<b>Cy-SNN</b>for sensing pH fluctuation under extremely acidic conditions. For the preparation of this probe, benzothiozolium moiety was chosen as lysosomal targeting unit and NIR fluorophore, and barbituric acid moiety was fused in the polymethine chain of probe to introduce protonation center. Surprisingly, on the basis of the balance of quaternary ammonium salts and free amines, the pk_avalue of<b>Cy-SNN</b>was calculated as low as 2.96, implying that<b>Cy-SNN</b>can be used in acidic conditions with pH < 4. Moreover,<b>Cy-SNN</b>exhibited highly selective response to H⁺over diverse analytes in real-time with dependable reversibility. Importantly,<b>Cy-SNN</b>can be used to specifically target lysosome, providing potential tools for monitoring the function of lysosome in autophagy process.</p>","PeriodicalId":18596,"journal":{"name":"Methods and Applications in Fluorescence","volume":"10 2","pages":""},"PeriodicalIF":3.2,"publicationDate":"2022-02-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39945152","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}