Basic procedure for setting up a timelapse experiment using
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GFP4
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Make sure the Oko-lab environmental chamber is set to the correct conditions for your experiment. In no particular order turn on the computer and the wall switch labeled "microscope devices". This turns on the following devices: microscope stand, motorised stage, light sources and camera. Login using your agendo username and password. Double-click the NIS-Elements icon on the desktop and enter your user credentials. When the driver selection window pops up, ensure the 'first driver' is set to 'Phasics', and the 'second driver' is set to 'ANDOR Neo/Zyla'. 'Enable Multi Camera' has to be ticked. Then click on 'OK'.
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Turn the pE100 off. Press the FL1 button on the remote control pad and select a filter position. Check the list of filters for the one you need. Turn on the pE-4000and adjust the brightness accordingly.
| Warning |
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| The excitation wavelength is determined by the microscope filter set used but it is also possible to change the balance of the UV, blue and green/yellow/red brightness of the pE4000 using buttons on the keypad and in the software. If you change the balance make sure that you use the same LED settings for any specimens that you want to compare. The pE4000 has 16 wavelength options to choose from. It is perhaps easier to use optical configurations within the software to get the settings you want. |
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Select the 'Zyla' tab in the bottom left of the screen. Press the L100 button to redirect the light to the camera. Click the fluorescence optical configuration you need in the OC Panel. Click the live preview button in the toolbar to get a live camera image. Click the pE-4000 Pad tab and adjust the brightness of the illumination if necessary. A red exclamation mark appears in the OC Panel if there are any changes. Click the arrow next to the optical configuration to save the new brightness (red arrow A below). In 'Zyla Settings' change the exposure, binning and gain as necessary. Changes to the exposure are automatically written to the optical configuration so you don’t need to click the arrow.
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