Multifocal multiphoton microscopy based on multianode photomultiplier tubes

Kim, Ki Hean; Buehler, Christof; Bahlmann, Karsten; Ragan, Timothy; Lee, Wei-Chung Allen; Nedivi, Elly; Heffer, Erica L.; Fantini, Sergio; So, Peter T. C.

doi:10.1364/oe.15.011658

Cited by 105 publications

(117 citation statements)

References 41 publications

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“…The majority of current single beam confocal applications use mirror galvanometers to achieve raster scanning of the focal spot [4,5,24,29]. A pair of orthogonal mirror galvanometers are used to tilt the trajectory of a light beam along two axes.…”

Section: Mirror Galvanometer Scanningmentioning

confidence: 99%

“…To take full advantage of a multiplexed confocal approach, the detection may be made by discrete single-point detectors, which offer better signal-to-noise performance and temporal resolution than a CCD array [24]. For these methods, a foci array is generated by a lenslet array or spatial light modulator, and raster scanning is performed by the entire array [25,26].…”

Section: Introductionmentioning

confidence: 99%

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High-speed multifocal array scanning using refractive window tilting

Tsikouras

Berman²,

Andrews

et al. 2015

Biomed. Opt. Express

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Confocal microscopy has several advantages over wide-field microscopy, such as out-of-focus light suppression, 3D sectioning, and compatibility with specialized detectors. While wide-field microscopy is a faster approach, multiplexed confocal schemes can be used to make confocal microscopy more suitable for high-throughput applications, such as high-content screening (HCS) commonly used in drug discovery. An increasingly powerful modality in HCS is fluorescence lifetime imaging microscopy (FLIM), which can be used to measure protein-protein interactions through Förster resonant energy transfer (FRET). FLIM-FRET for HCS combines the requirements of high throughput, high resolution and specialized time-resolving detectors, making it difficult to implement using wide-field and spinning disk confocal approaches. We developed a novel foci array scan method that can achieve uniform multiplex confocal acquisition using stationary lenslet arrays for high resolution and high throughput FLIM. Unlike traditional mirror galvanometers, which work in Fourier space between scan lenses, this scan method uses optical flats to steer a 2-dimension foci array through refraction. After integrating this scanning scheme in a multiplexing confocal FLIM system, we demonstrate it offers clear benefits over traditional mirror galvanometer scanners in scan linearity, uniformity, cost and complexity. 277-296 (2015). 27. T. Nielsen, M. Fricke, D. Hellweg, and P. Andresen, "High efficiency beam splitter for multifocal multiphoton microscopy," J.

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Section: Mirror Galvanometer Scanningmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

High-speed multifocal array scanning using refractive window tilting

Tsikouras

Berman²,

Andrews

et al. 2015

Biomed. Opt. Express

View full text Add to dashboard Cite

show abstract

“…A current drawback of multifocal multiphoton microscopy is lower signal-to-noise ratios at deeper regions. Kim et al (2007) examined the penetration depth by multifocal multiphoton microscopy with multichannel PMTs and succeeded in improving the signal-to-noise ratio, although the spatial resolution of multichannel PMTs is insufficient for fMCI (8ϫ8, H7546B, Hamamatsu Photonics, Hamamatsu, Japan). 143) Ji et al (2008) report that the splitting-laser-pulse method improves the signal-to-noise ratio and reduces photodamage and photobleaching.…”

Section: )mentioning

confidence: 99%

“…Kim et al (2007) examined the penetration depth by multifocal multiphoton microscopy with multichannel PMTs and succeeded in improving the signal-to-noise ratio, although the spatial resolution of multichannel PMTs is insufficient for fMCI (8ϫ8, H7546B, Hamamatsu Photonics, Hamamatsu, Japan). 143) Ji et al (2008) report that the splitting-laser-pulse method improves the signal-to-noise ratio and reduces photodamage and photobleaching. 144) Although multifocal multiphoton microscopy has not been applied to in vivo vertebrate species, it has been shown to work in in vivo insect brains 145) and acute mouse brain slices.…”

Section: )mentioning

confidence: 99%

Current Application and Technology of Functional Multineuron Calcium Imaging

Namiki

Ikegaya

2009

Biological & Pharmaceutical Bulletin

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Here we describe recent applications and technical advancements of functional multineuron calcium imaging (fMCI), which monitors the firing activity of more than a thousand neurons through their somatic Ca 2؉ signals. fMCI is used for analysis of various neural circuits under normal and pathological conditions. In vitro fMCI is made more sophisticated by using multipoint illumination and scanning technology with spinning-disk and low-laser-intensity imaging, electron-multiplying charge-coupled device cameras, etc. In vivo fMCI is still developing. We review optical technologies for fast scanning imaging, deep tissue imaging, and recording from moving animals.Key words calcium imaging; functional multineuron calcium imaging; large-scale recording; scanning; spinning disk Review January 2009 1 Biol. Pharm. Bull. 32(1) 1-9 (2009) © 2009 Pharmaceutical Society of Japan * To whom correspondence should be addressed. e-mail: ikegaya@mol.f.u-tokyo.ac.jp tation is removed with a small aperture called a pinhole which functions as a spatial filter 31) ; that is, light emitted from the focal spot efficiently passes through the pinhole, while the vast majority of emission light arising from the outside area is physically blocked. To acquire a two-dimensional or three-dimensional image of a specimen, the focal spot is moved with servomotor-controlled mirrors called Galvano-mirrors, servomotor-controlled stages, etc. Scanned emission light is detected by a photomultiplier tube (PMT). PMT detects a photon via a photoelectric effect and amplifies the signal 10 5 -10 8 times through multiple charged cathodes, in which an incoming photoelectron produces secondary electrons. The amplified electrons are converted into electric currents by an anode.Nipkow Spinning Disk Confocal Microscopy: Spinning Disk with Multi-Pinholes The second implementation is the use of multi-point illumination with many pinholes on a spinning plate termed a Nipkow disk. A rotating spinning disk has multiple pinholes through which laser lines embody multisite focal illumination and detection. The commercially available apparatus, Yokogawa's spinning disk, is further sophisticated. The system uses an additional spinning disk that contains a ca. 20000 microlens array for light convergence on the corresponding pinholes on a Nipkow disk.32) The light focusing also helps reduce scattering light, improving the signal-to-noise ratio. In addition, pinholes are strategically aligned in space to achieve uniform illumination over the field of view. 33,34) The latest version of Yokogawa's spinning disk, CSU-X1, is more improved in terms of the light use efficiency, about two-fold higher than the previous version, CSU-10. In CSU-X1, reduction of mechanical noise from its disk-driving part allows realization of 10000 revolutions per minute. Because the helical pinhole distribution covers the whole field of view every 30°of disk rotation, CSU-X1 enables the maximum sampling rate of 2000 Hz.Charge-Coupled Device Technology Spinning disk confocal microscopy simultaneously...

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“…There are several methods for approaching the generation of these foci, with various advantages and disadvantages. Common approaches to the problem involve the use of micro-lens arrays [2][3][4][5][6][7][8], etalons [9], diffractive optical elements [10], or cascading beam splitters [11][12][13][14][15]. Since these techniques utilize closely spaced or overlapping beams, it is important to use a spatially-resolved detector and temporally separate the arrival time of the beams in order to avoid interference of the excitation beams [4][5][6]12].…”

Section: Introductionmentioning

confidence: 99%

Remote focusing for programmable multi-layer differential multiphoton microscopy

Hoover¹,

Young²,

Chandler³

et al. 2010

Biomed. Opt. Express

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Abstract:We present the application of remote focusing to multiphoton laser scanning microscopy and utilize this technology to demonstrate simultaneous, programmable multi-layer imaging. Remote focusing is used to independently control the axial location of multiple focal planes that can be simultaneously imaged with single element detection. This facilitates volumetric multiphoton imaging in scattering specimens and can be practically scaled to a large number of focal planes. Further, it is demonstrated that the remote focusing control can be synchronized with the lateral scan directions, enabling imaging in orthogonal scan planes.

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Multifocal multiphoton microscopy based on multianode photomultiplier tubes

Cited by 105 publications

References 41 publications

High-speed multifocal array scanning using refractive window tilting

High-speed multifocal array scanning using refractive window tilting

Current Application and Technology of Functional Multineuron Calcium Imaging

Remote focusing for programmable multi-layer differential multiphoton microscopy

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