SerialEM settings

View: bin 1, 0.2 s exposure

Record: bin 1, 5s exposure

Here’s how I set up SerialEM data collection on the Krios

LMM.st 

Save navi

Align to marker 

Go to 205x mag

Close out of montage

New montage – MMM

Add squares

Then check the MMM acquire script

Acquire at items – MMM

Find constrast-y contamination, add point, go to low dose mode, go to XYZ & shift to marker 

Find square without features, set focus point. Change view to square, bin 8, 0.5s exposure & shrink. Once perfectly centred, save to buffer P.

Put objective aperture in, check stigmation. 

Astigmatism by CTF, coma-free by CTF, astigmatism by CTF again. 

View needs to be same as buffer P i.e. small & square 

Set multi-shot parameters.

Acquire at items! 

Use winSCP to copy gain ref to own workspace (although I think Sasha does this already now). 

e2proc2d.py gainref.dm3 newname.mrc —meanshrink 2 

Copy the new gain ref back to capture data

Using puTTy log onto taylorimages (cryoemiscool)

screen -D -RR

sudo semi.pl

Add notes if necessary

Run! 

Old exposure settings – 5.9 s, 0.074 sec/frame. 80 frames total. 

New exposure settings – 5s, 0.05s/frame (100 total). Total dose 70e- (0.7 e-/frame)

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