(A) Setting Up Your Environment to Run Amira
1. Amira can currently only be run from jane, so you will first need to ssh to jane:
$ ssh -Y jane.crbs.ucsd.edu
2. Enter the directory that contains your reconstruction. For example:
$ cd /ccdbprod/ccdbprod2/home/CCDB_DATA_USER.portal/CCDB_DATA_USER/acquisition/project_20191/microscopy_87042/reconstruction
3. Start Amira:
$ /ncmir/local.linux.amd64/Amira-5.4/bin/start
If you want, you can setup your login script to alias this command to the command 'amira'. To do this, add the following line to your .bashrc file by editing it in a text editor (vi, emacs, nano, etc.):
alias amira='/ncmir/local.linux.amd64/Amira-5.4/bin/start'
If you do this, the next time you use Amira, you will only have to type the command 'amira' rather than the full path.
(B) Preparing Volumes
Your reconstruction should already be in the MRC format if it was calculated using IMOD or TxBR. If your reconstruction is from a 4k x 4k camera, you should be able to load the full volume into memory on jane. However, if it is from the 8k x 8k camera or a montage, you may need to bin it so it will fit into memory.
1. If your volume needs to be binned, use the following command to bin by a factor of 2:
$ newstack -bin 2 basename.rec basename.2Dbin2.rec
2. To perform the maximum intensity projection (MIP) calculations, you need to have a volume with the contrast inverted such that electron dense regions appear white. This is opposite from what we are used to. Since the MIP calculations take a long time and the MIP rendering really slows down Amira, it is helpful to perform the calculations on a binned volume.
If you have already binned your volume in the above step, run the following command to invert the contrast:
$ newstack -scale 255,0 basename.2Dbin2.rec basename.2Dbin2.inverted.rec
If you haven't already binned your volume because it is from a 4k x 4k camera, run the following command to perform the binning and contrast inversion operations simultaneously:
$ newstack -bin 2 -scale 255,0 basename.rec basename.2Dbin2.inverted.rec
(C) Loading the Volumes into Amira
The Amira GUI is divided into four main frames. In the image below, I've given them names that I'll refer to for the rest of this tutorial:
1. In the Pool frame, click the green 'Open Data' button. Select your current directory from the upper pull-down menu (it should be the top choice), and select the first volume (basename.rec OR basename.2Dbin2.rec). Click the 'Load' button and select 'MRC Volume' in the 'Dialog' pop-up window. Click the 'OK' button.
Pool --> Open Data --> Select Data --> MRC Volume
Right click on button --> Display --> Voltex
Left click on Voltex button
Color table
Luminance/alpha
Colormap --> Edit --> Adjust to histogram, then adjust the min and max values accordingly
Alpha scale = 1
Texture mode: 2D
Downsample: 2 2 2
Apply
(click the image to view it at full resolution)