SEMC TF20, Руководство пользователя, Страница: 10 / 30

Last updated 10/29/2020
9
Cryo Data Collection with the DE20
The DE-20 camera system is a direct detection device (DDD) manufactured by Direct Electron. It has
high signal-to-noise ratio (SNR) with consistent performance across all spatial frequencies and TEM
magnifications. “Movie mode” proves high-speed acquisition of a continuous stream of frames (32 frames
per sec max, unbinned full-frame) with nearly 100% duty cycle (no dead time between frames). Increased
data quality compared to TVIPS thru drift correction, per-particle motion correction, radiation damage
compensation, post-acquisition exposure setting, etc. It has a 5k x 4k field of view (5120 x 3840) with a
6.4um pixel pitch. It is fully retractable and needs to be manually retracted and warmed when not in use,
and inserted and cooled when in use. The temperature changes make it so that the camera needs a new
dark and gain reference with each new session. The DE20 is sensitive to stray light (operate with window
covered and lights off). Suggested mags for EN/FA/FC = 50,000x, spot size 5, 3840 x 5120, bin1; HL =
5000x, spot size 5, 960 x 960, bin 4; SQ = 550x, spot size 5, 960 x 960 , bin 4; GR = 97x, spot size 5, 960
x 960 bin 4.
Dose should be 2.1 - 2.2 electrons/pixel/frame (no more than 2.5 otherwise clipping will occur). At
50,000X this corresponds to ~45 electrons/Å
2
for a 2 second exposure (1.54 Å/pixel).
1. After step 6 of F20 User Guide (starting your session) ...
2. Open DE20 software to insert and cool down camera (NOTE- can only open MicroManager or
Leginon client/server at one time); DE-20 camera control on monitor input DVI-1
a. Close Leginon client on camera computer
b.
Open “
DE server GUI (OLD)
” on camera computer
(this program is laggy; wait 30 sec)
c.
“Cool down” and “extend” camera (when not in use, DE20 should be retracted and
warmed); it takes ~ 5 min to cool down camera to -38 Celcius
i. See Temperature - Detector (Celcius) on the DE Server (Camera Settings)
ii. Wait for cool down before opening Leginon client (the DE server will stop refreshing and
updating temperature reading if Leginon client is open; temperature will not be affected)
d.
“do not autosave” = true
3. Open Leginon client on camera computer (DE20 DVI1), microscope computer (TF20-D394), and
main computer (TF20LEGINON)
a. In terminal, type betaleginon
4. Setup Leginon session settings:
a.
Select “create new session” → “next”
b. Select holder (TF20-cryo-holder_AMI-2)
c.
In session description, start with F20 and then type info about sample → “next”
d.
Select project → “next”
e.
Image directory (use default… “/gpfs/leginon/username”)
f.
Connect to clients= tf20-de20 & tf20- d394 → “next”
g.
C2 size = refer to the white board → “finish”
5.
Once Leginon has started up, open the “application”
a.
Run → Application → MSI -T2 (3.3)
i. Main = TF20Leginon
ii. Camera = tf20-de20