1,720,964 research outputs found
The BrightEyes-TTM: an open-source time-tagging module for fluorescence lifetime imaging microscopy applications
No full textThe aim of this Ph.D. work is to reason and show how an open-source multi-channel and standalone time-tagging device was developed, validated and used in combination with a new generation of single-photon array detectors to pursue super-resolved time-resolved fluorescence lifetime imaging measurements.
Within the compound of time-resolved fluorescence laser scanning microscopy (LSM) techniques, fluorescence lifetime imaging microscopy (FLIM) plays a relevant role in the life-sciences field, thanks to its ability of detecting functional changes within the cellular micro-environment.
The recent advancements in photon detection technologies, such as the introduction of asynchronous read-out single-photon avalanche diode (SPAD) array detectors, allow to image a fluorescent sample with spatial resolution below the diffraction limit, at the same time, yield the possibility of accessing the single-photon information content allowing for time-resolved FLIM measurements.
Thus, super-resolved FLIM experiments can be accomplished using SPAD array detectors in combination with pulsed laser sources and special data acquisition systems (DAQs), capable of handling a multiplicity of inputs and dealing with the single-photons readouts generated by SPAD array detectors.
Nowadays, the commercial market lacks a true standalone, multi-channel, single-board, time-tagging and affordable DAQ device specifically designed for super-resolved FLIM experiments. Moreover, in the scientific community, no-efforts have been placed yet in building a device that can compensate such absence. That is why, within this Ph.D. project, an open-source and low-cost device, the so-called BrightEyes-TTM (time tagging module), was developed and validated both for fluorescence lifetime and time-resolved measurements in general.
The BrightEyes-TTM belongs to a niche of DAQ devices called time-to-digital converters (TDCs). The field-gate programmable array (FPGA) technology was chosen for implementing the BrightEyes-TTM thanks to its reprogrammability and low cost features. The literature reports several different FPGA-based TDC architectures. Particularly, the differential delay-line TDC architecture turned out to be the most suitable for this Ph.D. project as it offers an optimal trade-off between temporal precision, temporal range, temporal resolution, dead-time, linearity, and FPGA resources, which are all crucial characteristics for a TDC device.
The goal of the project of pursuing a cost-effective and further-upgradable open-source time-tagging device was achieved as the BrigthEyes-TTM was developed and assembled using low-cost commercially available electronic development kits, thus allowing for the architecture to be easily reproduced. BrightEyes-TTM was deployed on a FPGA development board which was equipped with a USB 3.0 chip for communicating with a host-processing unit and a multi-input/output custom-built interface card for interconnecting the TTM with the outside world. Licence-free softwares were used for acquiring, reconstructing and analyzing the BrightEyes-TTM time-resolved data.
In order to characterize the BrightEyes-TTM performances and, at the same time, validate the developed multi-channel TDC architecture, the TTM was firstly tested on a bench and then integrated into a fluorescent LSM system. Yielding a 30 ps single-shot precision and linearity performances that allows to be employed for actual FLIM measurements, the BrightEyes-TTM, which also proved to acquire data from many channels in parallel, was ultimately used with a SPAD array detector to perform fluorescence imaging and spectroscopy on biological systems.
As output of the Ph.D. work, the BrightEyes-TTM was released on GitHub as a fully open-source project with two aims. The principal aim is to give to any microscopy and life science laboratory the possibility to implement and further develop single-photon-based time-resolved microscopy techniques. The second aim is to trigger the interest of the microscopy community, and establish the BrigthEyes-TTM as a new standard for single-photon FLSM and FLIM experiments
Fluorescence lifetime analysis (FLA) for the screening of healthcare nanoformulations: towards a compact and versatile device
No full textFluorescence lifetime analysis (FLA) has become essential for non-invasive, real-time analysis of a variegate realm of compounds and materials in scientific and industrial fields. Here we present a compact and versatile device designed to apply FLA to the screening of healthcare nanoformulations which contain a luminescent active principle. In the case study here: Doxil FLA enables to decode the supramolecular organization and stability of the luminescent active principle within the nanoformulation in a non-invasive, rapid (seconds), and cost-effective manner. Contrary to currently used methods (e.g. TEM, SEM, CryoEM, HPLC), this device performs the analysis in the natural solvent with no need for sample chemical manipulation or labelling. This advancement holds promise for enhancing research and quality control in pharmaceutical industries. Also, we envision in the near future application of this technology to evaluate drug entry into cells and to monitor the change of drug supramolecular organization upon contact with biological environments, such as biological fluids, cells, tissues
Fast and portable fluorescence lifetime analysis for early warning detection of micro- and nanoplastics in water
No full text: The presence of plastic fragments in aquatic environments, particularly at the micro- and nano-scale, has become a significant global concern. However, current detection methods are limited in their ability to reveal the presence of such particles in liquid samples. In this study, we propose the use of a fluorescence lifetime analysis system for the detection of micro- and nanoplastics in water. This approach relies on the inherent endogenous fluorescence of plastic materials and involves the collection of single photons emitted by plastic fragments upon exposure to a pulsed laser beam. Briefly, a pulsed laser beam (repetition frequency = 40 MHz) shines onto a sample solution, and the emitted light is filtered, collected, and used to trace the time distributions of the photons with high temporal resolution. Finally, the fluorescence lifetime was measured using fitting procedures and a phasor analysis. Phasor analysis is a fit-free method that allows the measurement of the fluorescence lifetime of a sample without any assumptions or prior knowledge of the sample decay pattern. The developed instrument was tested using fluorescence references and validated using unlabelled micro- and nano-scale particles. Our system successfully detected polystyrene particles in water, achieving a remarkable sensitivity with a detection limit of 0.01 mg/mL, without the need for sample pre-treatment or visual inspection. Although further studies are necessary to enhance the detection limit of the technique and distinguish between different plastic materials, this proof-of-concept study suggests the potential of the fluorescence lifetime-based approach as a rapid, robust, and cost-effective method for early warning detection and identification of plastic contaminants in aquatic environments
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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High Frequency Digital Frequency Domain Fluorescence Lifetime Imaging System
Full text linkA system and method is provided for improved fluorescence lifetime measurement. A high frequency digital heterodyning technique is described in which photon counting mode detectors are sampled at a rate slightly slower than a digitally pulsed excitation signal. A digital mixer produces a difference frequency which contains the same information than the high frequency signal. This lower frequency is still suitable for fluorescence lifetime measurements in microscopy. The digital algorithm provides phases and modulations of detected photons binning them into phase steps and time windows. The digital heterodynig algorithm was implemented in a very low cost FPGA (Field Programmable Gate Array)
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
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