Three‐dimensional microscopy migrates to the web with “PowerUp Your Microscope”

Bonetto, P.; Boccacci, Patrizia; Scarito, Marco; Davolio, Mirko; Epifani, Mattia; Vicidomini, Giuseppe; Tacchetti, Carlo; Ramoino, Paola; Usai, Cesare; Diaspro, Alberto

doi:10.1002/jemt.20074

Cited by 9 publications

(4 citation statements)

References 13 publications

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“…As a final general aspect, once the best quality image possible has been obtained then sophisticated mathematical algorithms can be applied to enhance those fine details and features of interest to the biological researcher and to improve the quality of data to be used for 3D modelling (Weinstein & Castleman, 1971;Diaspro et al 1990Diaspro et al , 2004cvan der Voort & Strasters, 1995 ;Boccacci & Bertero, 2001 ;Vermolen et al 2004). Recently, an image restoration web service has been established to obtain the best quality 3D dataset from wide-field, confocal or 2PE optically sectioned samples (Bonetto et al 2004).…”

Section: General Aspectsmentioning

confidence: 99%

“…Acquisition and visualization is completely computer controlled by dedicated open software (EZ2000 ; Coord, The Netherlands; http:// www.coord.nl). Image processing based on deconvolution algorithms is performed by means of a web-based package designed and implemented at the Laboratory for Advanced Microscopy, Bioimaging and Spectroscopy (LAMBS) (Diaspro et al 2002b ;Bonetto et al 2004).…”

Section: Example Of a Practical Realizationmentioning

confidence: 99%

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Two-photon fluorescence excitation and related techniques in biological microscopy

Diaspro

Chirico

Collini

2005

Quart. Rev. Biophys.

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279

217

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This review is concerned with two-photon excited fluorescence microscopy (2PE) and related techniques, which are probably the most important advance in optical microscopy of biological specimens since the introduction of confocal imaging. The advent of 2PE on the scene allowed the design and performance of many unimaginable biological studies from the single cell to the tissue level, and even to whole animals, at a resolution ranging from the classical hundreds of nanometres to the single molecule size. Moreover, 2PE enabled long-term imaging of in vivo biological specimens, image generation from deeper tissue depth, and higher signal-to-noise images compared to wide-field and confocal schemes. However, due to the fact that up to this time 2PE can only be considered to be in its infancy, the advantages over other techniques are still being evaluated. Here, after a brief historical introduction, we focus on the basic principles of 2PE including fluorescence correlation spectroscopy. The major advantages and drawbacks of 2PE-based experimental approaches are discussed and compared to the conventional single-photon excitation cases. In particular we deal with the fluorescence brightness of most used dyes and proteins under 2PE conditions, on the optical consequences of 2PE, and the saturation effects in 2PE that mostly limit the fluorescence output. A complete section is devoted to the discussion of 2PE of fluorescent probes. We then offer a description of the central experimental issues, namely: choice of microscope objectives, two-photon excitable dyes and fluorescent proteins, choice of laser sources, and effect of the optics on 2PE sensitivity. An inevitably partial, but vast, overview of the applications and a large and up-to-date bibliography terminate the review. As a conclusive comment, we believe that 2PE and related techniques can be considered as a mainstay of the modern biophysical research milieu and a bright perspective in optical microscopy.

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Section: General Aspectsmentioning

confidence: 99%

Section: Example Of a Practical Realizationmentioning

confidence: 99%

Two-photon fluorescence excitation and related techniques in biological microscopy

Diaspro

Chirico

Collini

2005

Quart. Rev. Biophys.

Self Cite

279

217

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“…Internet, database and information technologies have been applied to facilitate the confocal data administration and image processing (Carazo et al, 1996(Carazo et al, , 1998Carazo and Stelzer, 1999;Lindek et al, 1999;Pittet et al, 1999;Swedlow et al, 2003;Bonetto et al, 2004). Carazo et al (1996Carazo et al ( , 1998 introduced BioImage database which is an objectrelational database for multi-dimensional microscopic images of biological specimens available through the World Wide Web (WWW).…”

Section: Applications Of It In Cellular Imagingmentioning

confidence: 99%

“…OME can automatically manage images captured from all optical microscopes including confocal, wide-field, and multi-photon systems. Bonetto et al (2004) presented a software package, named "Power Up Your…”

Section: Applications Of It In Cellular Imagingmentioning

confidence: 99%

Processing, visualization and analysis of 3D confocal microscopic images

Guan¹

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Confocal laser scanning microscopes have been widely employed in biology research for many years. The advancement of computer technology enables interactive visualization and quantitative analysis of volumetric confocal image datasets thus giving deep insights into cellular structures. This thesis details the computer techniques developed by the author to facilitate 3D confocal image studies. The key contributions of this work are in (1) confocal image acquisition and management, (2) 3D image processing, and (3) Virtual Reality (VR) enhanced visualization and analysis of cellular structures. With a large amount of confocal images produced with microscopes for different biological studies, the administration of these image datasets become crucial. We develop a solution enabling convenient storage and management of confocal image data with a central image administration database. Furthermore, the integration of Internet techniques with the database makes it possible for online data access in a collaborative research environment. Raw confocal image data has various problems thus usually needs to be processed before visualization and quantitative analysis. A fast interpolation algorithm to produce

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Multiphoton Microscopy

Diaspro

Caorsi

Bianchini

et al. 2006

Wiley Encyclopedia of Biomedical Engineering

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Nowadays, optical microscopy has evolved into three‐dimensional (3‐D) (x‐y‐z) and four‐dimensional (4‐D) (x‐y‐z‐t) analyses allowing researchers to probe even deeper into the intricate mechanisms of living systems. Within this scenario, two‐photon and multiphoton excitation (TPE, MPE) microscopy are probably the most relevant advancement in fluorescence optical microscopy since the introduction of confocal imaging in the 1980s. MPE microscopy has a 3‐D intrinsic ability coupled with almost five other interesting capabilities, namely: (1) MPE greatly reduces photo‐interactions and allows imaging of living specimens on long time periods; (2) MPE operates in a high‐sensitivity background‐free acquisition scheme; (3) MPE microscopy can image turbid and thick specimens down to a depth of a few hundred micrometers; (4) MPE allows simultaneous excitation of different fluorescent molecules reducing colocalization errors; and (5) MPE can be used to prime photochemical reactions within a subfemtoliter volume inside solutions, cells, and tissues. Besides, MPE fluorescence microscopy is not only revolutionary in its ability to provide the above‐mentioned features, together with other practical advantages, but also in its elegance and effectiveness of application of quantum physics. The MPE story, more specifically TPE, dates back to 1931, and its roots are in the theory originally developed by Maria Göppert‐Mayer that brought, passing through the scanning nonlinear microscope developed by Colin Sheppard in Oxford at the end of the 1970s, to the revolutionary and successful results on biological samples obtained at the Watt Webb laboratories by Winfried Denk and colleagues at the beginning of the 1990s. MPE can be considered a young microscopical technique. In general, it is more convenient to consider TPE, for the sake of simplicity.

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Three‐dimensional microscopy migrates to the web with “PowerUp Your Microscope”

Cited by 9 publications

References 13 publications

Two-photon fluorescence excitation and related techniques in biological microscopy

Two-photon fluorescence excitation and related techniques in biological microscopy

Processing, visualization and analysis of 3D confocal microscopic images

Multiphoton Microscopy

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