1,720,991 research outputs found
MicroMos: an open source software tool to obtain high-resolution panoramic images of 2D cell cultures
No full textOBJECTIVE:
In order to perform significant analyses, the microscope’s user is typically interested in acquiring high-detailed images representing an entire histological sample or well containing cells. Due to the finite size of the camera’s field of view, the microscopist has to find the proper trade-off between higher magnification factor and extension of the observed area.
Here we present MicroMos version 3.0, an open-source tool for building mosaics by stitching together more partially overlapping images. The method proposed is based on visual information only and the mosaics are built by incrementally stitching couples of images. The radiance of the original sample is preserved by compensating the vignetting effect of each stitched image, and the mosaics obtained can be used for quantitative analyses.
By exploiting MicroMos we studied confluence and proliferation of mesenchymal stromal cells (MSC) adherent on OSPROLIFE (Eurocoating, Cirè-Pergine, Italy), a commercial biomaterial in the form of granules. In order to acquire statistically significant data we analysed mosaics built using MicroMos, implementing customized algorithms to segment the MSC and automatically estimate of the percentage of the area of the granules covered by cells.
MATERIALS AND METHODS:
To validate the proposed mosaicing method we performed several experiments under different working conditions. In particular, in order to assess the quality of the mosaics obtained using different warping models and tonal adjustments we used six different sets of images of histological samples and cell cultures. We aligned the images according to different registration models and computed several metrics to estimate which registration method performs as the best.
Then, to study confluence and proliferation of MSC adherent on OSPROLIFE granules, we prepared several samples each containing 50 mg of OSPROLIFE granules and a different number of MSC. Cell confluence was evaluated by adding 4 μM Calcein AM before acquiring sequence of partially overlapping images.
RESULTS:
In our experiments we tested different registration approaches, confirming quite unexpectedly that the translational model does not always act as the best, although the motion of the microscope’ sample holder is apparently translational. Indeed, the sample holder could be slightly inclined, hence yielding non-negligibly affine, or even projective, transformations between subsequent images.
CONCLUSION:
MicroMos version 3.0 is freely distributed as an open source tool, endowed with a graphical user interface, at the website: https://sourceforge.net/p/micromos/. Its usability makes building mosaics of microscope images at subpixel accuracy easier. Furthermore, optional parameters for building mosaics according to different strategies make MicroMos an easy and reliable tool to compare different registration approaches, warping models and tonal corrections. Finally, by analysing mosaics of granules at different time intervals we also obtained quantitative data regarding cell proliferation, confirming that MSC adhere onto the OSPROLIFE granules and proliferate over time
Over time homogeneity and stability of mesenchymal stromal cells 3D spheroids built using base-level laboratory equipment
No full textMesenchymal Stromal Cells (MSC) are widely used for tissue engineering applications because of their capacity to promote tissue regeneration while modulating immune responses. Three-dimensional (3D) MSC aggregates, also known as multicellular spheroids, better preserve phenotype and innate properties with respect to cells cultured in monolayer. They are the preferred format for many cell therapy and tissue engineering applications, and for bioprinting uses.
Many approaches have been proposed in literature to generate spheroids composed of cancer cells, and most of them can be applied when constructing MSC spheroids. Efficiency, cost, and size of the produced spheroids are typically considered to compare different generation approaches. The pellet culture method provides a cost-effective and extremely rapid method to generate 3D MSC spheroids. Only a benchtop centrifuge and sterile polypropylene conical tubes, typically available in every biological laboratory, are required. However, although the pellet culture method is widely used to generate MSC spheroids, few studies have been performed to analyse the morpho-biological homogeneity and stability during time in culture of the obtained spheroids.
In this study we monitored for two months changes in size and morphology of a set of spheroids generated by the pellet culture method. Each spheroid was imaged with brightfield microscopy twice a week. AnaSP (open-source software freely available at: http://sourceforge.net/p/anasp) was used to compute volume, sphericity and jagging (i.e. indentation degree of the spheroid's border) of each spheroid over time.
The volume of the MSC spheroids showed a decreasing trend. However, just fifteen days from spheroid generation, all the values remained constant and similar, demonstrating good volume stability and homogeneity after an initial settling time. Sphericity of the MSC spheroids kept always high values, thus indicating that the shape is generally close to a perfect sphere. Jagging degree values were always very low, proving that MSC spheroids are characterized by a compact border. We also performed confocal and histological analyses to better investigate their stability. Hematoxylin&Eosin stained cryosections were analysed at different time points to evaluate the internal cellular architecture. Then, the cell viability and distribution on the spheroid surface was evaluated with Live&Dead staining and confocal imaging. No significant changes in internal distribution and spheroid compactness were observed up to 30 days from the spheroids generation.
In conclusion, we can assume that the MSC spheroids generated are homogenous and stable for a long time interval. Accordingly, the proposed enhanced pellet culture method can be used as a reference system for laboratories interested in easily obtaining, without need for specialized equipment, stable homogeneous populations of MSC spheroids to be used in long term studies and applications
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
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
In vitro quantitative analysis of mesenchymal stromal cells migration towards tumours
No full textAnalysis of in vitro migration of cell populations plays a key role in studying a wide range of dynamic cell behaviours. Cell migration is dependent on a multitude of signals ranging from growth factors to chemokines secreted by injured cells and/or respondent immune cells. In the last decade, several studies were addressed to investigate tropism of Mesenchymal Stromal Cells (MSC) for some types of cancers. Understanding the factors involved in regulating MSC migration towards tumours is essential to ultimately develop novel clinical strategies aiming at using MSC as vehicles to deliver antitumor drugs. However, the mechanism behind MSC migration is still in its infancy, and the main cause is the lack of quantitative methods to analyse the MSC motion.
The analysis of objects motion relies on image-based tracking techniques and several methods have been recently proposed to track cells in vitro. However the vast majority of them are limited to fluorescently labelled cells. Very few tracking tools can analyze images taken using label-free microscopy techniques (e.g. phase contrast), that do not compromise the biological status of the cell and are the most common ways to observe living cells. However, to accurately track irregular-shaped, flat, and poor contrasted cells, such as MSC observed in phase-contrast, is really challenging. Furthermore, the majority of the tracking tools require relevant image processing skills, which strongly limits their practical usefulness within the biologist community. User-friendly and versatile open source software with track editing possibilities would boost live cell analysis research. In this work we introduced CellTracker, a user-friendly open-source software tool for tracking cells, including MSC. CellTracker is freely available at: http://celltracker.website/ and it works with Windows, Macintosh, and UNIX-based systems.
By using CellTracker and μ-Slide Chemotaxis2D (IBIDI, Madison, Wisconsin, USA) we performed experiments to analyse the motility of MSC when seeded near to cancer cells, simply seeding MSC in a chamber communicating to another one containing conditioned medium from MG-63 osteosarcoma cells. As negative and chemotaxis controls, we used respectively fresh culture medium and culture medium supplemented with serum.
From the CellTracker analysis we could not detect any significant change in MSC motility towards the culture medium with or without serum, whereas a different behaviour was observed by using culture conditioned medium taken from flasks previously containing MG-63 cells. We can therefore argue that MSC migration towards tumour cells is mediated by the presence of specific proteins (e.g. chemokines) secreted by cancer cells in the culture medium. These preliminary results pave the way for future applications such as the possibility to develop hybrid disposable diagnostic devices where biological fluids (e.g. blood) sampled from the area of interest are analysed by using the MSC motility as a cancer sensor
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
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