Taking Pictures-Zen 2011
TAKING PICTURES ON THE ZEISS TISSUE CULTURE
MICROSCOPE
MICROSCOPE START-UP
If needed, turn on the Fluorescent Light Source.
Turn on the microscope.
Turn on the computer.
Power Button
Log in with your WVU credentials – select Zen2011
software.
Eyes/Camera Switch
GETTING STARTED
Place your sample on the stage and focus.
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If you are imaging in brightfield, choose the correct Phase settings based upon the
objective you are using.
If you are imaging using fluorescence, change the filter by pushing the filter bar under
the stage to either the left or the right.
Once you have your sample in focus, switch the light path from the eyes to the camera.
CAMERA SETTINGS
Click on the Live Button. This will display your image
in the Live window. The Live button will turn into a
spinning red circle with Stop in the center.
In the Dimensions tab, you can change the
Camera Mode.
Greyscale will give you the highest sensitivity for
fluorescence.
Color works best for non-fluorescent samples.
Taking Pictures with Zen 2011
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Under the Microscope Components tab, select the
objective you are currently using. This will provide the
proper scaling settings to your image.
Under the Camera tab, you can set the Camera
Exposure Time.
You can manually change the exposure time using the
slider or by typing in a particular time. The Auto
Exposure button will let the computer choose the best
exposure time for your sample.
For color images, adjust the White Balance to make
the background white. If you use the Pick… button,
you will select an area on your image that should be
white. If you want the software to set the white
balance, find an empty field (something that should be
completely white) and choose Auto.
Display Settings
Select the Display tab to see a histogram display of image intensities.
Use the buttons at the top and bottom of the histogram to change how the image is displayed
on the monitor. This does not change the data.
Choose Gamma 0.45 for the best color reproduction. Use Automatic Min/Max to find the
optimal setting for contrast and brightness.
Range Indicator will highlight saturated pixels.
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CAPTURE IMAGE
Click on Snap to acquire an image. After you snap the image, a Graphics tab appears. You can
then add graphics to the image:
Scale bar
Scale Bar:
Arrow
Shapes
Spline Tool
Make sure you have the correct objective selected under the Microscope
Components Heading.
Select the Scale Bar icon. Draw the scale bar on your image. To change the color
of the scale bar, right click on the scale bar and select Format Graphical
Elements. Then choose the color/font/line thickness you want.
Spline Tool:
The spline tool allows you to outline regions of interest, and then provides
information on the region, including area and intensity mean value*.
*Click on the Measure tab to get the chart
pictured below.
*You must burn-in the annotations to get them to stay on the saved image. Go to Graphics →
Burn-In Annotations. Once you burn in the annotations, you cannot remove them from the
picture.
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SAVE IMAGE
Go to File, Save As to save your image.
The *.czi format saves the image and header information. This format contains the most
information and will allow you to make changes or additions in the future. You can get a free
copy of Zen 2011 so you can open these files on your computer.
The TIFF file format saves just the image. This is a general file format that can be opened in
other programs such as Photoshop and Powerpoint. When you save as a tiff, make note of the
checkboxes at the bottom.
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USING THE SPOT METER FUNCTION
If you are having trouble focusing your sample on a low power
objective, you can use the Spot Meter to help you find the perfect
focus point.
Bring your sample into focus (or as close as you can get), and select
the Spot Meter box. A red dash box will appear in your live image
window. You can change the position and size of the box.
Right click inside the box, and from the menu that
appears, select Focus Bar.
The red line in the Focus Bar indicates where the
perfect focus position for that image selection should
be. Try to get the blue box to fill to the red line.
SHADING CORRECTION
If you have uneven illumination in your sample, you can use shading correction function to fix
this.
Move to a blank portion of your slide that is clear of any tissue or debris. You may need to
move the image out of focus to achieve this.
Under Post Processing, Select the box for Shading Correction.
The background image is now saved and applied directly to your live image.
*Note:
The shading correction only works if you have a background image that is free of
ALL debris and flaws. If you have any specks or debris, they will show up as
shadows on the final image.
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