CM120 Operation Instruction¶
Aug 8, 2016
Oct 3, 2021
This document tries to list steps and procedures for a typical daily operation on CM120. You can use it as guidebook to help you when you are sitting with CM120, especially if you are a new user. I here assume you are already familiar with scope interface, what the knobs and buttons do etc.. You might use the section titles as quick bullet p ints, but the explanation inside each sections are supposed to be useful and informative.
If you have suggestion how to improve this document to make it more useful, please feel free to let me know. Thank you!
all the turning knobs on the panel and push buttons are marked like
Check log book to see if there have been any problems¶
It is always a good idea to check the LOG book. You can find useful information listed there, such as:
If there have been any recent problems with the scope.
The conditions that were used last, especially the filament saturation.
Any special note that last user wants you to know.
Check vacuum status¶
The status must be Ready before you can operate the scope. If not, you should ask for help and report it to the manager. Usually, if the vacuum is not ready, it is due to one of the following reasons:
The scope is malfunctioning.
The air pressure is not within a good range(the building compressed air may be down?).
The cooling water is off.
Vacuum being Ready means:
Top line of vacuum page shows ready.
P3 < 50, IGP < 26 (normally, they are shown 0, 5).
LEDs for UVAC and HiVAC are lit.
Add Liquid Nitrogen to the BIG anti-contaminator dewar¶
LN2 in the cold trap dewar is necessary for fast vacuum recovery. It is very useful, especially when you need to change grids and/or do a cryo session. However, if you are working with a negative stain or plastic section - a dry grid, the scope can still run without LN2 cold trap. Unless you are running automatic, long overnight session, you should always use it.
Generally, when you first put LN2 dewar to its stand, you want to be SLOW, or the strong evaporation will make LN2 spilling.
Apply High Tension¶
Turn High Tension (H.T.) on if it is off by pressing the
H.T. button on
the panel. From
Parameter page, set it to 120kV or the voltage you
Turn filament on¶
It is recommended to turn the filament on while on the
page where the actual filament current number is shown and the
Check saturation and gun tilt, and then saturate the filament¶
The procedure is as following:
Desaturate the filament about 2-3 clicks.
multi-function X,Yto get the best shape for the tip and maximum intensity as well.
Bring the filament current to saturation.
Check C2 aperture mechanical position and C2 stigmatism¶
Steps to adjust the C2 aperture mechanical position are:
Make sure the C2 aperture is in.
At around 5kX, cross the beam first and center beam using track ball on the left panel, then spread the beam until its diameter is close to 5cm ring on the screen. Adjust mechanical
screws X,Yon C2 aperture to center the beam.
Repeat this process 2-3 times.
At high mag ~30kX or higher, change the
Intensityof beam through the crossover and adjust the
screws X,Yto make the beam symmetric when spreading out.
There are two ways to check and adjust the C2 stigmatism:
When the filament is desaturated, press
Stigbutton and select
Cond, then adjust
Multi-function X,Yso that the details in the filament shadowing image can be clearest and sharpest.
When changing the beam intensity, use the
Multi-function X,Yto adjust the beam into symmetric and round shape, i.e. not elliptical.
For this purpose, don’t pay attention to the beam shape when beam is exactly at cross point, as that more reflects the shape of the crystal tip, rather than stigmatism. You want to make beam round when spreat out.
Check specimen holder & load grid¶
This is important. If you see any problem with any of the holders, report it to the manager immediately. Otherwise, you could be the one held accountable for the damage.
Several details about the holder must be checked carefully before use:
Overall shape is good, and there is not obvious damage.
Make sure there is no crack or any other damage on the O-ring. If you do see a damage, like a cut etc., ask manager to replace it for you. Check if there any dirt or fibril on the O-ring. You might want to clean it gently with alcohol and slightly re-grease it. Do not over-grease. The main function of the grease is to lubricate.
Gently secure the specimen grid on its position. Use the tool pin to open and close the clamping device.
Insert specimen holder into column¶
Be careful! Only at this stage, you might damage the scope or specimen holder mechanically. Be sure that you understand what you are doing. Should you feel any confusion about this procedure, please stop and ask for help.
For the sake of filament crystal, it is REQUIRED to turn down the filament to 10 before inserting the specimen rod. That way, in case IGP shoots high, there will be no subtancial damage to the LaB6 tip crystal. In general, filament should be kept at 10 or completely off until IGP recovers to below 26.
The procedure to insert a room temperature specimen rod is below:
define airlock pumping time as 60 seconds, from Vacuum - Cryo page.
Reset stage tilt angle to 0 if it is not.
Insert rod in, with the Pin at 3’oclock position.
As soon as it reach the end, rotate rod CLOCKWISE with some pushing force so that the pin slides into the locking groove at 5’oclock position. You should feel the rod goes “in” about 8mm.
Wait until the red LED on the stage disappears. Dismiss the “non-standard” flushing message on the screen by pressing
Resetbutton at lower left corner of the screen display.
Turn rod Count-Clockwise until pin is at 12’oclock position, while watching IGP reading. You should adjust your rotating speed to keep IGP < 40.
Adjust the specimen height to the eucentric height¶
Eucentricity is a fixed reference point in a scope. It is the intercross point of stage axis and column axis. We want to observe our specimen grid at this height level so that the actual magnification doesn’t differ much from day to day. And scope is designed to perform better when specimen is at such height. It is good to have the habit to always adjust specimen to eucentric height after rod insertion. On CM120, the procedure is as following:
Have beam seen on large screen, at ~3000X, and find a feature on the grid.
Rotate stage back and forth by pressing
Adjust joystick Z to minimize the shifting of the feature.
Check Beam Tilt Pivot Point X, Y¶
The prerequisites for Pivot-Point is specimen being at eucentric height and objective being preceisly at focus level. The order is important here.
Make sure the specimen is at eucentric height.
Take out Obj. aperture.
Beam Tilt Pivot X.
Merge image feature by adjusting
Focusknob. This is to precisely focus the image.
Merge beam using
Repeat the last two steps for Beam Tilt Pivot Y.
Alignbutton again to exit.
Check voltage and current rotation centers¶
This step is to align the beam to make it parallel to the axis of the column. The purpose of this step is to make beam to hit specimen perpendicularly. A coma is not a good thing, as it generates some phase error to the data.
The procedure is below.
Alignbutton and select
Currentfrom the same page.
Multi-function Knobsto let the wobbling be symmetrical around the center of the beam. The feature at very center of the large screen has minimum shift.
If you perform this with Obj aperture in, then re-check the centering of objective aperture.
The step size button on
Focus is used to control the amplitude of the
The steps used here only give “roughly” parallel beam to the axis. If you need very accurate “0” tilt beam, a different alignment procedure - Coma-Free is needed.
Put in objective aperture and center it¶
It is important to know what size of the obj. aperture you are using. You don’t want to use too small size to actual cut off useful high resolution signal. Meantime, you don’t want to use the aperture size too large, as the non-usable high resolution beam becomes noise to your image. This reduces signal to noise ration unnecessarily.
The position of the aperture could affect the obj. lens stigmatism. Therefore, you want to do this step before you finally check Obj lens stigmatism.
Here are the steps to insert and center Objective lens aperture:
Make sure the large screen is down, to prevent CCD from damage.
Switch to diffraction mode by pressing the diffraction
Adjust camera length to ~1m using magnification knob.
Defocusknobs to see the shape edge of the obj. aperture.
Adjust the related mechanical screws on aperture holder to choose the proper size of the aperture and center it to the central beam on diffraction pattern.
Switch back to image mode by pressing
Check Objective Lens stigmatism¶
The obj. stigmatism should be corrected as much as possible, and it should be checked for every netative stain low-dose image that you are taking, as staining material might change field in local area. This is a bit hard by hand. Even with lastest version of SerialEW, this can be done by software, it is still not easy and time efficient. However, slight stigmated image is not critically bad, as it can be corrected as part of CTF correction computationally.
Here are steps to correct Obj lens stigmatism, manually:
Go to a relatively high mag., such as 100,000X, and focus the image.
If possible, acquire continous CCD image with live FFT so Thon rings can be seen.
Obj, and select proper stepsize.
Adjust the stigmatism using
Multi-function X,Yuntil it becomes minimum at all defocus levels. (It shows up more at close to focus.)
Magnification setup for three modes¶
Here are some typical magnificaton setup for Low-Dose condition.
One might use diffraction mode to
Search, which usually gives better
contrast but the “image” might be distorted. The final magnification depends
on target pixelsize on image. If possible, use the same magnification for
Record to eliminate the defocus offset between the
two magnifications. The off-axis distance is usually about 1.5 - 2.0
Align an identified area under
This step is to insure that what you see under low mag. (Search mode) will be the same area you get under imaging mag. Here is how I do it:
At Exposure mode, MECHANICALLY drive an identified spot to the center of the screen.
At Search mode(and usually in Diffraction mode also), using the
Multi-function knobsto backtrack the identified spot at the center of the screen (electronically). This uses Image Shift or Diffraction Shift (when Search mode is set up in Diff mode) to “shift” image without actually moving the stage position.
You can use a corner of a mesh as the identified spot for a negative stain specimet or to use an ice burn mark in the cryo case.
When you are done with your session, perform finishing up procedure.
Specimen rod out.
Reset Stage Position, X, Y, Z and A.
Cryo-cycle, normally for 2-3 hours.
Data display OFF.
Log your session on logbook.