Leginon

Every acquisition node has a pause button in the tool bar, shaped like what you see on your VCR. The question is which one to use. The pause actually occurs right before the next data acquisition in that node. See the section on pausing in the trouble shooting chapter. I used general description there so it might be confusing for a specific situation. Here are some specific situations:

1. In MSI non-queuing mode, pausing at Hole node is most commonly done.

2. In MSI queuing at exposure targeting and after all hole images are taken, you can no longer pause at Hole node, obviouisly. In this case, pause at Exposure node (see also the chapter for MSI and the section on Queuing Example 1: Exposure Targeting).

Drift manager will also check for drift and wait until drift stop before moving on. Therefore, you can continue (clicking the play button at the same node where you paused) immediately after the fill and let drift manager to decide when to acquire the next final exposure.

Anchi

Be aware that the pause buttons in Leginon do not usually pause instantly. If it is in the middle of acquiring several holes on a square, and you click pause, it will have to finish acquiring the current hole (and maybe all the exposures on that hole) before it will actually pause. So after you click pause, wait until the "?" icon appears next the node you paused, then it is safe to fill up your N2.

Also, letting Drift Manager determine when drifting has stopped is very effective, but watch out for cases where filling the N2 might cause such a fast drift, that consecutive images acquired by Drift Manager do not even correlate with each other (sometimes resulting in a false measurement)

Jim

The topic/ thread on automated top up makes me realise I'm not sure if I'm doing the correct thing in manual top up of liquid nitrogen. How soon do you normally restart Leginon (by pressing the play button in Hole or Exposure nodes - after the previous pause with the pause button)? Do you normally restart immediately which you suggest is possible below or after a few minutes (or half an hour which at least one of the local microscopists says they do when working manually without Leginon)?

William

"anchi" wrote: Every acquisition node has a pause button in the tool bar, shaped like what you see on your VCR. The question is which one to use. The pause actually occurs right before the next data acquisition in that node. See the section on pausing in the trouble shooting chapter. I used general description there so it might be confusing for a specific situation. Here are some specific situations:

1. In MSI non-queuing mode, pausing at Hole node is most commonly done.
2. In MSI queuing at exposure targeting and after all hole images are taken, you can no longer pause at Hole node, obviouisly. In this case, pause at Exposure node (see also the chapter for MSI and the section on Queuing Example 1: Exposure Targeting).

Drift manager will also check for drift and wait until drift stop before moving on. Therefore, you can continue (clicking the play button at the same node where you paused) immediately after the fill and let drift manager to decide when to acquire the next final exposure.

Anchi

Depending on the person, some starts immediately, others prefer to wait for 5-10 min. The false measurement Jim talked about only happens in the first minute or two, especially if you have not filled for a long time. In addition, this rare event would only affect one exposure sequence because new drift check will be made for the next exposure and focus sequence as long as you do activate the drift check function in your autofocusing.

Those who wait do so because they see no reason to expose the grid to the electron (although only in fa preset and at the focus position) when it is obvious that the grid would drift and because they are not in a hurry of going somewhere. It is true that if the drift is large and last for a long time (happens only if you let the cryo-stage dry out before refill), the region that is exposed would streak out and might get close to your final exposure target since the correction of targeting is made only after the drift has stopped.

Automated top up is a very different situation because Norhof at the moment is completely independent. Norhof does not know that Leginon is taking pictures and Leginon does not know that Norhof is filling the stage. By pausing Leginon when you manually fill the stage and the cold trap, you keep Leginon from acquiring images during the fill.

Anchi

By the way, when doing manual LN2 top up (and pausing Leginon) is it advisable to press the "Declare Drift" button in the Drift Manager node before unpausing by pressing the play button in either the Exposure or Hole nodes (whichever was paused before) or is this unnecessary or even likely to introduce problems?

William

Declare drift should not cause problem unless the target it need to check comes from an image that can no longer acquired properly. An example of

this would be that the target came from an objective aperture removed

grid image but the user is now collecting data with the aperture in.

"Declare Drift" event is sent automatically in MSI at several places already.

These include when you correct defocus by moving z with the goniometer, after the drift settles if "check drift" is turned on in autofocusing sequence and a drift is detected by the process. If you turn on "Declare drift when queue submitted", drift will of course declared, too, there. Unless you have

targeting problem from these, declare drift manually after nitrogen filled should only add benefit if any.

Node Description Chapter of the full Leginon manual has a description of the drift manager. It explains further how it behaves.

Anchi

I should clarify what I mean by "Declare drift after nitrogen filling only add benefit if ANY". In general "Declare Drift" before unpausing for nitrogen filling add benefit by forcing a check on the targeting and is recommended. It does not add additional benefit if other processes also send the drift time stamp in the same time frame. An example would be that when you unpause in Focus node before it initiates drift check of its own and the stage is not yet settle. You would get a "Drift Declared" from the drift checking any way.

Anchi