Zeiss LSM 880 Operating Manual page 383

LSM 880
View 1 is the master view and cannot be moved during the process, the button View 1 is therefore
inactive and grayed out. Any of the other views can be activated by pressing the according button (e.g.
View2, Fig. 528). The activated view is automatically displayed in the image (the associated channel is
turend on). To move the view relatively to View 1 the sliders and spin boxes can be used; X for the x-axis,
Y for the y-axis, Z for the z-axis and Angle for a roation. The axis that is not present in the maximum
intensity projection is grayed out and inactive (Z for the Front View and X for the Side View).
Addintionally to the rotation Angle, the position of the rotation point can be defined. This is done by
positioning the red Cross overlay within the maximum intensity projection image window to the desired
position, the view should be rotated around (Fig. 530).
The Reset button moves all views back to the position they were at, as the maximum intensity projection
was generated.
Each view should be activated once with its corresponding button and positioned, resulting in a matching
overlay to View 1. Afterwards the maximum intensity projection is switched to either Front View or
Side View; displayed in the same image window.
Again all views must be moved using the according sliders Y, Z and Angle to produce a perfect overlay
to View 1.
Switching from Front View to Side View should be repeated at least twice, to adust the positioning of
the views.
To controll if the manual alignment is satisfactory, the Preview button produces a new image of the
channel and the time point, which is registered with the provided settings. The Preview image will
contain fused views, when Mean Fusion has been selected at the Fusion Method drop down list.
If the result is satisfactory, one can proceed with the remaining settings for Multiview Processing.
If the data was acquired with dual side illumination, which has not yet been fused, the method to fuse
both illumination sides is available in the Dual Side Fusion drop-down menu (Mean Fusion, DFT
Fusion, Maximum Fusion).
Mean Fusion: The average intensity for each pixel is calculated using the values of both illumination
sides.
DFT Fusion (Discrete Fourier Transform Fusion): The Fusion is performed based upon the published
method ("Multiple imaging axis microscopy improves resolution for thick-sample applications", Jim
Swoger, Jan Huisken, and Ernst H. K. Stelzer, OPTICS LETTERS / Vol. 28, No. 18 / September 15, 2003).
Fine structures and details are more visible after the fusion, which might be worsened when using Mean
Fusion. However, this process can create noticeable artifacts in the resulting image.
Maximum Fusion: The intensity of each pixel of both illumination images is compared and the value
with the higher intensity is used for the resulting image.
For Mean Fusion and Maximum Fusion the checkbox Fusion Subset in X with Preview can be
activated (Fig. 531). With these controls it is possible to define which area of the left and the right
illumination image should be used for fusion.
When this function is chosen, the image is displayed in the Preview Tab in the image container
(Fig. 532). In this preview mode the left hand image shows the original dataset and the right hand image
the preview of processing. Using the dimension sliders within the image control area the Z-Position,
Illumination side and View can be chosen.
In Fusion Subset in X with Preview the currently chosen View is displayed in the Current View box.
With the Left slider or input box the area of the left illumination side is chosen. This is marked at the
slider itself with a red line, the input box shows the pixel value along the y axis. Furthermore the cut off is
shown in the preview image with a red line for easy orientation. It is best to set the illumination slider
of the image control area to 1, showing the left side illumination of the original data.
10/2014 V_01
CHAPTER 1 - SYSTEM OPERATION
Left Tool Area and Hardware Control Tools
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