Olympus IX71 microscope
Hamamatsu camera
Sutter excitation filter wheels
Uniblitz transmitted light shutter
Metamorph 5.0 software for camera integrated with shutters & filters

overview picture

Always set condenser for Kohler illumination.
1. Focus on sample
2. Close Field Diaphagm
3. Move condenser up and down using knob on right until center octagon is in focus
4. Open field diaphragm to edge of visible field, not all the way.

 

Objective Lens Condenser setting
10X Ph1 for phase contrast
10X BF for Brightfield
40X UCL

Ph2 for phase contrast
40X UCL BF for brightfield
60X oil BF for brightfield

Recommend NOT using the 60X air or 100X oil lenses.

 

Important for best image quality:

The 40X lens has an adjustable collar. You turn this ring to optimize the lens for the type of chamber you are imaging. A number 1.5 coverslip needs the ring adjusted to the 0.17 position. A plastic flask or dish will need a setting in the 1 to 2 range. (The units aremm.)

However, you cannot easily see the collar underneath the stage. Therefore:
1. Focus on sample best possible.
2. Reach under stage to turn ring on objective a little.
3. Refocus, turn ring, and repeat until image has maximum crispness.

Also, this may also be done when in the fluorescence mode. It is important to make this adjustment or else your pictures will not be in proper focus.

 

Fluorescence:

Color Fluorophore Filter turret Sutter electronic filter position
Green (FITC, GFP, YFP, Alexa488, Cy2, etc.) 1 1 or 0
Red (Rhodamine, Alexa 568, RFP, mCherry, etc.) 2 or 3? 2
Green and red together (by eye only; not with camera) 2 0
Blue (Dapi) 4 0
This table needs to be checked and completed; please let me know if you have info.    

 

Computer control of microscope

All equipment on before starting software.

Uniblitz box set to   A   N.C.   Remote

In software Devices > Illumination pulldown menu for Transmitted Light or fluorescence.

Use either Journal Recorder or Multi Dimensional Acquire menu to set up protocols to take transmitted and fluorescence images.

 

 

handwritten notes

another photo of electronics controls

 

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comments, questions, suggestions for this web page: Michael.Cammer@med.nyu.edu or mcammer@gmail.com