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Hyperspectral microscopy system

PARISS® Hyperspectral Microscopy


The PARISS Model “PHSI” operates in field scanning mode in darkfield scatter, fluorescence, luminescence, %transmission, %reflection and absorption.

PARISS hyperspectral microscopy work flow

Hyperspectral microscopy work flow (Details)

PARISS hyperspectral microscope model: PHSI

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  • Prism based imaging spectrometer for extended wavelength range and light throughput (prism vs diffraction grating details)
  • Mounting interface: “C” mount to most research upright and inverted microscopes
  • Wavelength range: 360 to 920-nm simultaneously
  • Spectral resolution: ~1-nm at 436-nm
  • PARISS imaging spectrometer (Details)
  • Scientific CCD/CMOS spectrum detector
  • Observed image scientific CCD/CMOS
  • Data processor
  • Computer operated microscope stage
  • Custom hyperspectral microscopy software. See here
  • Includes a MIDL calibration lamp to validate wavelength accuracy
  • Optional: NIST certified light source.
  • Optional Xenon illuminator for darkfield nanoparticle characterization
  • Calibration standards: Available MIDL wavelength calibration lamp and a “SYLPH”  NIST certified radiometric light source.
  • Sold either as an accessory to an existing microscope or as a complete system, contact LightForm.

How Hyperspectral Microscopy Works

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Hyperspectral microscopy correlates spectra presented by the field of view (FOV) with spectra in a reference  spectral library (RSL).  An unlimited FOV is acquired by placing a sample on a slide mounted on a computer controlled translation stage on a microscope. The sample is illuminated with white light for darkfield scatter, %transmission or %reflection, or with a laser for fluorescence or other excited states. To see a video that describes how the PARISS hyperspectral microscope works click here.

Each object in the field of view will present a spectrum.  If there are tens of thousands of objects, then PARISS will acquire tens of thousands of spectra many of which are common. The challenge associated with handling thousands of unique and often common spectra is handled with custom software that sorts and classifies all spectra in the FOV.

All or some classes of spectra can then be entered into an RSL. Each library spectrum can be pseudo-colored and linked to a unique target “spectral object”.  The presence of one or more pseudo-colors then confirms the location and presence of a target object.

Future samples can then be scanned, and the spectra presented by all objects in the FOV will be correlated with those in the library.  Those that meet a minimum correlation coefficient will then be “painted” onto a gray scale image. Various data processing options including counting correlated objects and various mathematical functions.

To see a video that describes how the PARISS microscope works in darkfield click here.

Creating Reference Spectral Libraries

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The PARISS hyperspectral microscope acquires many thousands of spectra over an unlimited field of view (FOV).  The PARISS software evaluates all spectra and sorts them into “classes.”  One class will likely be “background spectra,” the remaining spectra will be target objects.

Some, or all, classes of spectra can be added into an RSL.  Each library spectrum will be pseudo colored either by the software or the instrument user.

All, or selected spectra from the FOV that correlate with a library spectrum, will then be “painted” with pixel-perfect accuracy onto a grayscale image  of the FOV.

Hyperspectral Microscopy FAQ

Do I need a new microscope to set up a hyperspectral microscopy system?

No, not necessarily. If you have an existing research quality microscope with a video port, you are already in very good condition. LightForm supplies hyperspectral microscope modules that interface with most microscopes. Contact LightForm for more information.

Can a modular hyperspectral microscope be updated with time?

Yes. The PARISS hyperspectral microscopy system is designed for researchers on a tight budget to start slow and build the system up with time.

Does my hyperspectral microscope need additional illumination for fluorescence emission?

Yes, your microscope must have a fluorescence excitation source such as mercury, or an LED lamp. You will also need long-pass emission filter cubes.

Does my hyperspectral microscope need additional illumination for darkfield scatter for nanoparticle characterization?

Yes, most likely. Ideally, you need a Xenon light source to provide the wavelength range and signal strength for effective darkfield scatter spectroscopy.

Does a hyperspectral microscope need a translation stage?

Yes, if you plan on scanning an unlimited field-of-view such as a sample on a microscope slide. LightForm can help.

Do I need special hyperspectral microscope software?

Yes, hyperspectral imaging produces tens of thousands of spectra in a short period. PARISS software is written in PYTHON and includes all the functions you need to create a hyperspectral microscopy image. The PARISS software includes spectroscopic classification, reference spectral libraries, math functions, and stage control.

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Hyperspectral microscope image of nanoparticles internalized in cells

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Reference spectral library

The PARISS hyperspectral microscope software generates a reference spectral library (RSL.)

Hyperspectral microscope image of multi-fluorescent labeled tissue

Hyperspectral microscopy used to delineate multiple overlapping fluorophores in a tissue section

How Hyperspectral Microscopy Works

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