Leginon

Please be more specific on how multiple image shift targeting doesn't work. Does it not taking images, or does it targeted wrong or does it cause drift ?

We use multiple image shift targeting all the time. The accuracy is worse as your shift goes up to 2-3 um range but within a hole it is quite accurate.

Stage Postion targeting always causes more drift, just as you have observed.

There are a few things that can go wrong in image shift targeting, but I will wait for your reply before I elaborate.

Hi Anchi,

Thanks for the quick reply.
The size of the hole we used is 4um, we tried to set 3 targets in a hole, since we limit our Illumination Area to around 1.3um.
So the distance among target could be around 1.7um
For some reason, it just took 3 exposure on the same position.
We're checking the Image Shift Matrix calibration.
Anything else could go wrong?
One more info, we're using K2 super-res for exposure.

So it is is a targeting accuracy problem.

A few things to check:

1. How good is your K2 Counting calibrations ?

• You should check both image shift matrix and stage position matrix at the mag of your parent image (hl preset) and of the final image using Navigation node.

• Also see these:
  • Troubles_with_Targeting
  • Scaling_Stage_Position_Matrix_Calibration

2. How did you get super-res calibrations ?

It is close to impossible to get calibration with super-res. Therefore, we scale them from Counting mode. Dig you follow this? K2_calibration

If,after checking these, it still behaves wrongly, I will need you to send me your calibration matrix.

Hi Anchi,

Thanks.
Our K2 counting calibration is fine, though it seems the super-res calibration isn't.
We'll try counted first, and make sure everything work, then we'll switch to super-res (We'll follow your recommendation).