HOLOSCOPE

What is HOLOSCOPE?

HOLOSCOPE is a fully digital holographic system that allows you to perform two-photon functional imaging. HOLOSCOPE HOLOSCOPE enables the implementation of a patented parallel laser scanning two-photon imaging with fewer optical components compared to conventional two-photon microscopes and no moving parts thus improving mechanical stability.
HOLOSCOPE requires an infrared femtosecond pulsed laser source and can be combined with a high-speed camera (We give full support in selecting the best cost effective components for laser and camera).

Holoscope Software allows to illuminate the desired intensity pattern by calculating the phase shift for every pixel of the SLM. Phase masks are generated through iterative algorithms guaranteeing that each point of interest is illuminated by a diffraction-limited focal volume. Software example Software sums to the obtained phase mask a Fresnel lens pattern to avoid undiffracted light on the sample. Software allows to generate full frame two-photon images at variable frame rate and resolution that can be interactively selected. Software allows the user to select an arbitrary number of points of interest directly chosen from the acquired image and to generate an hologram illuminating each of them with diffraction-limited focal points.

On Neurophotonics!

HOLOSCOPE results reached the cover of SPIE Neurophotonics Journal in March 2015. Neurophotonics: http://neurophotonics.spiedigitallibrary.org/article.aspx?articleid=2119120

HOLOSCOPE Specifications

Download HOLOSCOPE Brochure
  • SLM chip size: 792 x 600 px
  • SLM pitch size: 20 um or 12.5 um
  • Holographic focus shift range: 30 um
  • Hologram calculation time: around 30 s
  • Input digital communication: DVI-D port
  • Control software: for Microsoft OS
  • Power supply: EU/US standards

HOLOSCOPE Applications

Calcium imaging

Holoscope allowed to monitor the activity of hundreds of neurons simultaneously in brain slice preparations thus proving to be capable of mapping the functional impact of synaptic activity in intact neuronal microcircuits with single cell and kHz resolution.

Neuropharmacology

Holoscope may help to understand how neuromodulatory systems control neuronal and microcircuit functions. It is possible to syudy the effects of neuromodulators (acetylcholine, serotonine, dopamine, noradrenaline) and drugs on neurons.

Photostimulation

Photostimulation is the use of light to artificially activate biological compounds, cells, tissues or even whole organisms. In particular photostimulation may be used for the mapping of neuronal connections between different areas of the brain by "uncaging" signaling biomolecules with light. Photostimulation uses light to uncage a compound that then becomes biochemically active, binding to a downstream effector. For example, uncaging glutamate is useful for finding excitatory connections between neurons, since the uncaged glutamate mimics the natural synaptic activity of one neuron impinging upon another. To this aim, Holoscope can be employed to selectively and precisely stimulate the release of caged compounds so that activating synaptic pathways.

Optogenetics

Optogenetics is the combination of genetics and optics to control well-defined events within specific cells of living tissue. It includes the discovery and insertion into cells of genes that confer light responsiveness; it also includes the associated technologies for delivering light to cells of interest, and for assessing specific readouts, or effects, of this optical control. By virtue of the arbitrary selection of the illumination points in the sample, HOLOSCOPE represents an optimal tool for optogentics.
What excites neuroscientists about optogenetics is control over defined events within defined cell types at defined times, a level of precision that is most likely crucial to biological understanding even beyond neuroscience. The significance of any event in a cell has full meaning only in the context of the other events occurring around it in the rest of the tissue. Even a shift of a few milliseconds in the timing of a neuron's firing, for example, can sometimes completely reverse the effect of its signal on the rest of the nervous system. And millisecond-scale timing precision within behaving mammals has been essential for key insights into both normal brain function and into clinical problems such as parkinsonism.

Fluorescence Correlation spectroscopy

FCS (Fluorescence Correlation Spectroscopy) is a correlation analysis of fluctuations of the fluorescence intensity. One application consists in the analysis of the concentration fluctuations of fluorescent particles (molecules) in solution. In this application, the fluorescence emitted in a very tiny space in solution containing a small number of fluorescent particles (molecules) is observed. FCS is such a sensitive analytical tool because it observes a small number of molecules (nanomolar to picomolar concentrations) in a small volume (femtoliter). Furthermore FCS enables the observation of fluorescence-tagged molecules in the biochemical pathways in intact living cells. Holoscope is the ideal experimental setup for FCS experiments variants involving multiple excitation volumes, by acquiring simultaneously millisecond scale signals from multiple confocal volumes. Holoscope can perform the following FCS experiments:
  • Pair correlation spectroscopy, in order to map the barriers and obstacles to diffusion inside the field of view.
  • Dual spot cross correlation spectroscopy, in order to map high speed flows such as blood circulation in vessels.
  • Spatio temporal cross correlation spectroscopy, revealing preferential directions in the diffusion of underresolved fluorescent particle.

Neuroscience

Both Human Brain Project (HBP) and BRAIN Initiative (Brain Research through Advancing Innovative Neurotechnologies also referred to as the Brain Activity Map Project) require high throughput screening technology to assess their ambitious goals, emulating the human brain capabilities and mapping the activity of every neuron in the human brain respectively.In the last decades the methods employed to monitor the activity of neuronal circuits have exponentially increased.

Dentrite blue

Examples of Holoscope results and software analysis

A section of mouse intestine (final resolution 1 micrometer/pixel)
Comparison of mouse kidney obtained with Holoscope (left) and with a conventional confocal microscope (right). Cerebellar granule cells with neuritic prolongations. Neuronal and astrocitic membranes coloured with a selective marker (ANNINE6-plus). Cerebellar slice bulk loaded with FURA2-AM, in which several granule cells and a large Purkinje cell with its dendritic arborization are visible. Optical resolution of Holoscope obtained through fluorescent polystirene beads
Dinamics

Quotations and Questions

If you need a quotation or any information please send an email to info@holobe.eu or fill the following template. THANKS!

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