Olympus FV3000 Veiligheidshandboek - Pagina 11

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There's no general rule: these adjustments mainly depend on your sample.

4.6 Image Adjustments – Setting the Gain and Offset properly

Check image saturation level by clicking

Hi-Lo icon. The image will

change color to show red and

blue pixels. Blue indicates that the signal

in a given pixel is black, with a value of 0.

Red indicates that the signal in a given

pixel is saturated (over-exposed) with a

value of 4095.

For optimal performance, the image

needs to be so that then entire image is

set to the limits of the dynamic range.

Different sets of experiments may have

different fluorescence intensities.

Adjust the image using a combination of

the excitation Laser power, HV and Offset:

Reduce either the HV or the laser power,

until red pixels disappear. Increase Offset until a dusting of blue pixels in the background are

observed.

Click LSM start to capture an image.

4.7 How to do a Z stack for a 3D Image (xyz)

1. Under Z Series: ON for Z.

2. Select Live 2x or Live 4x to see the

specimen and click Register under origin

tab to save your focal plane where your

image is focused. After completing the Z-

stack. Move can be selected to move back

to the focal plane of focus.

3. Turn the focus knob clockwise and move all

the way to the top of your sample. Click

4. Turn the focus knob anticlockwise and move

all the way to the bottom of your sample.

Click Register under end tab.

5. With system optimized selected, the

program will determine the optimal settings

for your Z-stack. You can override these

settings using the slices or z-step size boxes.

Beware that changing these will alter the z-

resolution you attain.

Last updated on Mar 2022