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  • Optical Projection Tomography (OPT)

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  1. On the OPT reconstruction computer, open the Nrecon folder on the Desktop
  2. Double click the GPUReconServer icon and then the Nrecon application icon
  3. Press the open folder icon at the top left of the Nrecon page
  4. Find the folder with your files, double click any of the 1199 tif files 
  5. Adjust the position of the green line across the sample (for embryos, this would be across the heart)
  6. In the Reconstruction tab on the top right of the screen, select the Settings tab
    • adjust the misalignment compensation value until the sample in the viewing window appears in focus
    • adjust the Ring artifact reduction value to 4
  7. Back at the Start tab, press the Fine tuning box
    • select Post-alignment option and select the following parameters:
      • Number of trials: 5
      • Parameter step: 1
    • Press Start button
    • Use the up and down arrows at the top of the screen to select the best image (this should be the one that looks most focused)
  8. Repeat step 7 above but change the Parameter step size to 0.5
    • Again, use the up and down arrows to select the best focused image
  9. Note: If you are unable to find a focused image it may be that your sample moved during the scan. In this case, you will have to rescan your sample
  10. Once you are satisfied with your misalignment compensation setting, press the Add to batch button
  11. Repeat for each of your samples
  12. Once all your samples have been added to the batch, press Start batch on the bottom left of the screen
  13. Once the batch is completed, you will find the reconstructed tif files in your sample folder
  14. These will have to be transferred to your hpf directory (the OPT reconstruction computer has very little available memory to store images)

Step 3 - Producing a 3D mnc file 

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