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3D stability for imaging

US Patent Number 9,019,363

  What is Nano-Cyte®?


Features
  • 3D stabilization up to 3 nanometers
  • Active positional control over days
  • Corrects for temperature gradients and drift
  • Simultaneous image acquisition and stabilization
  • Particle localization analysis
  • Particle position rendering
The Nano-Cyte® single molecule imaging system eliminates the microscope drift that limits advanced fluorescence imaging methods. With Nano-Cyte® you no longer need to be concerned with temperature gradients, sample drift, and microscope drift. Unprecedented stability in the nanometer regime allows long term experiments as never before.

The Nano-Cyte® works by using the image of fluorescent fiduciary references, sparsely distributed within the sample, to localize these emitters in all three dimensions. The Nano-Cyte® uses this 3D localization information to provide active position adjustments to the sample, thus eliminating drift in the experiment.

Nano-Cyte® is the complete stabilization and image acquisition instrument for advanced fluorescence microscopy. Our integrated approach to 3D stabilization yields image stability up to 3nm in X,Y and Z axes. Nano-Cyte® has proven stability over days and is a unique offering that promises to revolutionize advanced microscopy methods.

  Hardware



The Nano-Cyte® is comprised of a high performance three axis piezo nanopositioning system coupled with a two axis motorized micropositioning stage. These precision motion capabilities enable the active positional control and particle tracking features of the Nano-Cyte®. The nanopositioner is a flexure guided piezoactuated design with integrated PicoQ® sensors for absolute position sensing and nanometer precision under closed loop control. The micropositioning stage enables the user to have a large range of travel for surveying samples prior to engaging the active stabilization. All motion devices are controlled by the Nano-Cyte® controller via USB 2.0 interface.

The Nano-Cyte® hardware is compatible with Mad City Labs RM21™ open microscopy platform and most models of inverted optical microscopes.

  Method


Nano-Cyte feedback control diagram
Video of Nano-Cyte three dimensional image-based stability for live cell imaging
Video: Nano-Cyte®: 3D Image Stabilization System
The Nano-Cyte® feedback control system. Stabilization is based on the imaging pathway.

  Software


The native Nano-Cyte® software performs 6 important functions:

  • Stabilization
  • Image acquisition
  • Device control
  • Particle localization analysis
  • Rendering of particle position
  • Tracking over multiple fields-of-view (FOV)

The Nano-Cyte® 3D stabilization occurs simultaneously with image acquistiion and incorporates reference selection, reference localization, and calibration statistics. Acquired images are saved in TIFF format and can be exported to ImageJ and other 3rd party software for post-acquisition processing.

Nano-Cyte® device control ensures precision motion control and the ability to incorporate a variety of external user devices such as EMCCD cameras, shutters and light sources. Compatibility with LabVIEW™ and μManager facilitates even greater user device control and flexibility.

Post-acquisition features of Nano-Cyte® enable the localization of particles within an image and the three dimensional rendering of particle positions.

Nano-Cyte® is compatible with LabVIEW™, μManager, ImageJVI and rapidSTORM. In addition, Nano-Cyte® has an exportable DLL to allow wider functionality with 3rd party software platforms.

  Stability Data


Nanometer Stability over Days


Stability data measured over 44 hours for the Z-axis. The red line indicates the measured drift. The blue line indicates the stabilized position when using Nano-Cyte®. Similar results were observed simultaneously for the X and Y-axis. These data below demonstrate the efficacy of the Nano-Cyte® under non-monotonic drift conditions.

Nano-Cyte Z axis stability data

Effective Stability Using Nano-Cyte®


The Nano-Cyte® data below shows position changes performed on the z-axis over a period 1 hour. Similar results were observed simultaneously for the X and Y-axis. These data demonstrate that the Nano-Cyte® effective stability is 3nm.

Nano-Cyte Z axis bit size step data

  Nano-Cyte® Specifications


Stability (X, Y, Z-Axis) 3 nm
Drift rate compensation (typical) 10 nm/sec
Stabilization rate (typical) 1 frame/sec
Maximum data acquisition rate Camera dependent
Nanopositioning range of motion 200 μm × 200 μm × 200 μm
      Position noise (total) 0.4 nm
      Step size 3 nm
Micropositioning range of motion 25 mm x 25 mm
      Encoder resolution 20 nm
      Minimum step size 95 nm
Controller Nano-Cyte®
Communication USB 2.0
TTL outputs 4 channels
Output image format TIFF
Software Nano-Cyte®
Software compatibility LabVIEW
μManager
ImageJ & ImageJVI
Exportable DLL
RapidSTORM
Microscope compatibility Nikon Ti, TE Series
Zeiss Axio Series
Olympus IX Series
RM21™
Supported EMCCD Camera types Andor
Photometrics
Hamamatsu
Power supply 90-260 VAC (50/60Hz)
Operating system Windows Vista/7/8



Additional Information

Nano-Cyte® Brochure
Nano-Cyte brochure

Nano-Cyte® Video
Video of Nano-Cyte three dimensional image-based stability for live cell imaging
Technical notes about sample preparation and applications are available by request.

References


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mclgen@madcitylabs.com       phone: 608.298.0855       fax: 608.298.9525

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