Michael Hooker Microscopy Facility (MHMF)

Leica MX16FA Fluorescence Stereo Microscope/Macroscope

0.  Notices:

Due to a color camera failure on the Leica inverted widefield Color camera (MicroPublisher) is being shared with the Leica Inverted widefield on the other side of the curtain.

1.  The Scope:

• Basic stereo scope
• Combi attachment
• Objectives
  • 0.63x
  • 1.0x
  • 2.0x
  • 5.0x Combi (Monocular)
• Illumination
  • Incident - Schott halogen lamp with adjustable fiber optic pair
  • Transmitter - Rotterman illuminator in base
  • Epi-Fluorescence - 120W or (Prior 200W HBO metal halide)
• Fluorescence filters -
  • Dapi, Alexa350
  • FITC/CY2/GFP/Aelxa488/etc.
  • Rhodamine/TexasRed/CY3/Alexa568/Alexa594/etc.
  • CY5/Alexa633/Aelxa647/Draq5 and more
  • Leica Designations:
    • UV
    • CFP - Ex 436/20 nm Em 480/40 nm
    • CY5 - Ex 620/60 nm Em 700/75 nm
    • dsRed - Ex 545/30 nm Em 620/60 nm
    • GFP2 - Ex 480/40 nm Em 510 LP
    • GFP3 - Ex 470/40 nm Em 525/50 nm
    • 2x Transmitted (blocks Metal Halide)
    • Do not have
      • TexasRed (use dsRed)
      • YFP (use GFP2 long pass or GP3 band pass if orange/red autofluorescence is significant)
    • Combi dichroics
      • CFP
      • GPF3
      • CY5
      • dsRed
      • Triple
• Color camera - MicroPublisher 5.0 MPixels (2560 x 1900) light split 50/50 between camera and right eye piece
• Orca ER B/W CCD camera (sometimes B/W camera - Cool Snap HQ2 1300x1030 pixels) Light manually switchable 50/50 camera/left eye piece or 100% camera & 0% left eye piece)
• Computer - WindowsXP on MHmicroscopy domain, 2.67 GHz CPU, 2.0 GByte RAM, Disk storage 299 GBytes
• Acquisition software: C-Imaging 6.6.0.19
• External connections - For thumb drives and other portable devices
  • USB
  • Firewire (1394) - use port from mother board (not the interface cards connected to the cameras)
• Foot Pedal with programmable functions - zoom or focus or fluorescence filter changer (usualy not installed)
• Motorized focus, zoom, filter changer, shutters, which are both under manual or computer control
• Additional stereo image recording capability with dual camera acquisition is being developed.  Stereoptic computer viewing will be possible.

2.  Using:

• Contact Michael or Neal
• Power on computer and monitor (usually left on 24/7)
• Login with MHmicroscopy account
• Turn on Hamamatsu Orca ER b/w camera (not necessary if only using the color camera)
• Ensure MicroPublisher is present and Firewire cable is plugged in (not required if only using b/w camera)
• Turn on switch on the back of the base of the scope body.
• Turn on switch on top of focus drive
• Start C-Imaging v 6.6.0.19
• Selected camera
• Ensure cooling is enabled is using the OrcaER
• Selected 8 bit imaging (generally)

• For Transmitted light:
  • Illumination from base
  • Illumination from above with Schott lamp
    • K

• For Fluorescence

3.  Access:

• Contact Michael or Neal

4.  Local File Storage

• Drive C:\Users \\Gould\users1 Usable space <30 GBytes

• Drive D:\

  \\Gould\users2

  Usable space 179 GBytes

• DVD reader and writer - DVD/CD creation and burning software is Roxio Creator 9 Silver

5.  Off Line Analysis

• Image viewing, processing and quantification functionality present on this scopes computer (\\Gould) is also available on several image processing workstations in the same room as the scope.  These computers include \\Schott, \\Brewster, \\Nomarski, \\Snell.  Data may be accessed from the \\Gould computer as  \\gould\users1  or  \\gould\users2  from these image processing workstations even while the scope is being used by others for image acquisition. 
• Deconvolution and basic 3D rendering is available in the C-Imaging installation on the computer called \\Nomarski and using Volocity on various computers. 
• Additional 3D rendering and time lapse processing, analysis and quantification is available on the image processing work station \\Zernike (located in the adjacent room 6033a)
• De-skewing software will be investigated if there is a demand.
• (Some) available functionality - time lapse viewing - exporting of time lapse and z-series data as .avi movie files, tiff file series - extended focus generation - 
• Available on \\Snell, \\Nomarski,  \\Zernike QuickTime-Pro for .avi assembly from .tif series and conversion to/from .mov (quicktime) formats.

6.  Notes:

The USB connector on the back of the scope body sometimes comes loose.  When loose can not control scope with the computer and the Leica MZ16 tab in C-Imaging capture will not be able to read the filters installed.

7.  Links:

• Leica Stereo Scopes
• http://www.stereomicroscopy.com/website/sc_sms.nsf/(allids)/6b1bdedb34a25503c12570610047ddbd

8.  Acknowledgements:

• Funding for this resource from the North Carolina Biotechnology Center is greatly appreciated.

		Copyright 2001-14  Dr. M. Chua, Office of Research, University of North Carolina, Chapel Hill, NC 27599
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