Dear GA users,
Has anyone had a problem where you already zeroed Z on a tile, you move the objective and return to the same tile and Z is no longer focused? I've been having this problem recently where after zeroing Z on lane 4 column 2 row 25 and then proceeding to align X and the XY drift and going back to that tile to check Z, and it will no longer be in focus. I used to be able to have Z still in focus or at least it is only off by very little (~100) but now, it is always off by at least 1000-3000. I also have problems going through the fine focusing procedure as the lab tracking sheets are set up for (by 100's) because the Z keeps going off by increments in the 1000's.
Also, when I try to align X and the XY drift, between that and rechecking my Z, I often have to reset my XY drift by the exact same increment as when I zeroed it before going back to check if Z is still in focus as if the origin was never set the first time. And my X and XY drift alignments are set in greater values than before. I used to be able to set X = 3 to maybe 15 but now it's more like 30-55. To me, these differences seem too large. Are your X values going up to the 50's? More troubling is of course that even with a bead flowcell, I have to constantly reset the XY drift by the same increment as if the origin was not set.
The problem is once I quickly do the fine focus on Z and go straight to first base incorporation, then it will usually look ok and this problem does not reoccur consistently. However, we have had our previous run fail because the Z suddenly went out of focus in the middle of the run. We could see that image A started to get blurry and C went completely out of focus. My hypothesis is something is lose in our machine, but with the inconsistency of reoccurrence, it has been very difficult for us to receive service on our machine that regains our confidence in subsequent runs.
Does anyone else have a similar experience and do you have any idea what is wrong and how can we fix this?
Thanks,
PF
Has anyone had a problem where you already zeroed Z on a tile, you move the objective and return to the same tile and Z is no longer focused? I've been having this problem recently where after zeroing Z on lane 4 column 2 row 25 and then proceeding to align X and the XY drift and going back to that tile to check Z, and it will no longer be in focus. I used to be able to have Z still in focus or at least it is only off by very little (~100) but now, it is always off by at least 1000-3000. I also have problems going through the fine focusing procedure as the lab tracking sheets are set up for (by 100's) because the Z keeps going off by increments in the 1000's.
Also, when I try to align X and the XY drift, between that and rechecking my Z, I often have to reset my XY drift by the exact same increment as when I zeroed it before going back to check if Z is still in focus as if the origin was never set the first time. And my X and XY drift alignments are set in greater values than before. I used to be able to set X = 3 to maybe 15 but now it's more like 30-55. To me, these differences seem too large. Are your X values going up to the 50's? More troubling is of course that even with a bead flowcell, I have to constantly reset the XY drift by the same increment as if the origin was not set.
The problem is once I quickly do the fine focus on Z and go straight to first base incorporation, then it will usually look ok and this problem does not reoccur consistently. However, we have had our previous run fail because the Z suddenly went out of focus in the middle of the run. We could see that image A started to get blurry and C went completely out of focus. My hypothesis is something is lose in our machine, but with the inconsistency of reoccurrence, it has been very difficult for us to receive service on our machine that regains our confidence in subsequent runs.
Does anyone else have a similar experience and do you have any idea what is wrong and how can we fix this?
Thanks,
PF
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