Export of Transition List and Method for Sciex 6500+ broken

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Export of Transition List and Method for Sciex 6500+ broken Will Thompson  2020-08-19
 

Hi Brendan et al

Export of transition lists and of methods for Sciex 6500+ is not currently working in Skyline -daily or 20.x build, meaning method development is not possible on this instrument. Specifically this is the case for negative ion small molecule data. Some problems may be specific to that use case, but others seem generic. Not sure how long this has been going on, we have not done any routine method development on this system in a while. There are a number of issues. Recognized so far:

  1. Skyline will not allow a negative collision energy or declustering potential. These are required for negative ion acquisition for Sciex. When you paste in a negative CE or DxP using the import or Ctrl+V method, Skyline does not give an error, but it just puts nothing in the appropriate fields.
  2. Even if you change the polarity of the CE in Skyline to (the incorrect) positive value, then you export an acquisition method, the method contains a random negative number instead of the CE that was programmed.
  3. Even if you change the polarity of the CE in Skyline to (the incorrect) positive value, then you export a transition list, the TL is the incorrect format and Analyst cannot import it.

I hope you can reproduce these errors with the attached documents but if not we can set up a Zoom call for me to walk through in more detail. Please let me know if you need more information.

Cheers

Will

 
 
Brian Pratt responded:  2020-08-19

Hi Will,

Thanks for the report, I'm investigating now. We'll let you know if there are any questions.

Brian

 
Brian Pratt responded:  2020-08-19

Hi Will,

Looking at the Skyline code dating back 5 or more years ago there are a lot of checks to explicitly avoid negative CE values. Probably that's just an artifact of Skyline's early days where negative ion mode wasn't supported. Has this in fact ever worked for you in negative ion mode? I just want to make sure that we're dealing with an obsolete assumption rather than a newly broken output.

Thanks

Brian

 
Will Thompson responded:  2020-08-19

My guess is that this never worked. The small amount of real method development I have done on this machine has been manual. The Sciex 6500+ is a new machine for us. Since Sciex requires negative CE and DxP values for its methods, my guess is that negative ion method export and method development options (CE optimization, DxP optimiziation, etc) doesn't work for Sciex? It would be strange that this is the case, if nobody has ever reported it. But if it is the case, this will be an important feature upgrade for small molecules!

Cheers

Will

 
Brian Pratt responded:  2020-08-19

I think it must be that no one else has tried this.

Do you agree that the proper,more general rule that it makes no sense to emit CE and DxP values of polarity that does not agree with the machine's ion mode? That's effectively what's happening now for positive ion mode.

Thanks

Brian

 
Will Thompson responded:  2020-08-20

Hey Brian

Unfortunately, not. Waters uses positive CE and cone voltages etc for all ionization modes, as does i believe Thermo. With Sciex the CE and DxP voltages match the analysis polarity. Not sure about Agilent or Shimadzu. I'm not sure I would implement a 'check' for the relationship between ionization mode and CE polarity unless you can somehow make it vendor-specific...which of course you will not typically specify in the document before you paste the transition list. This is probably going to be a case where Skyline will need to just allow either positive or negative values (or zero) in these fields in order to make sure the proper format can be exported. It will be possible for the user to make a 'mistake' with the value of their collision energy, but i'm not sure Skyline can help that unless some much more advanced logic is involved at the export method/TL step.

Cheers

W

 
Brian Pratt responded:  2020-08-20

I think you're right, but we can at least do the vendor-appropriate thing during export. We'll get on that.

Thanks for being such an active member of the Skyline community!

Brian

 
Brian Pratt responded:  2020-08-20

Hi Will,

Might it make more sense for Skyline to always deal with (i.e. display and calculate) CE and DxP as positive values, and simply format them as negative values on output when appropriate to the ionization mode? Currently by specifying negative explicit CE values you are implicitly excluding the use of most mass spec types, not to mention ionization modes. Seems like Skyline should just magically deal with that.

Thanks,

Brian

 
Will Thompson responded:  2020-08-20

Perhaps...but it may also cause confusion when a publication lists collision energies for Sciex as a negative value, then the user must know that they need to change them to positive for Skyline to understand them, then they will be changed back when skyline exports a method or transition list for sciex machine? Seems like if Skyline does not allow negative CE values then it would require some sophisticated 'audit/error logic' where Skyline will tell the user what they need to do to make the CEs be recognized (if they paste in negative CE values, for instance), and why the program is either A) going to change them to positive, or B) going to require the user to do it.

At the risk of making it even more complicated...the whole issue of negative CE values also will potentially create an issue for CE optimization workflows, whereby an 'increase' in CE would actually require a subtraction from the value (making it more negative) rather than more positive. Makes my head hurt. Maybe Skyline always just converting everything to a positive value, even when pasted negative, is possible? Then it is swapped to negative upon export for the instruments that require it?

W

 
Brian Pratt responded:  2020-08-20

Maybe Skyline always just converting everything to a positive value, even when pasted negative, is possible? Then it is swapped to negative upon export for the instruments that require it?

That's what I was thinking, yes. We'd use the existing CE optimization code that builds the ramp as postive values, but write those as negative values for Sciex negative ionization.

 
Brian Pratt responded:  2020-08-20

Skyline will tell the user what they need to do to make the CEs be recognized (if they paste in negative CE values, for instance), and why the program is either A) going to change them to positive, or B) going to require the user to do it.

Yes, I would imagine Skyline issuing an error that CE values must be presented as positive values only, but will be written as negative when appropriate to the hardware and ionization mode.

 
Brian Pratt responded:  2020-08-21

One more question, Will - should Compensation Voltage receive the same treatment?

 
Will Thompson responded:  2020-08-21

HI Brian

It looks like that for negative ion mode, there should be negative voltages for all of the settings. Abbreviations used in the literature vary, but CE, DxP, CxP, and EP are all negative. It might be a good idea to follow up with your specific Sciex contact to be sure about all the export expectations. If you don't have one, I would recommend Kenzie Pearson (Mackenzie.Pearson@sciex.com), one of Sciex excellent apps chemists in the small molecule space.

Cheers,

W

 
Brian Pratt responded:  2020-08-21

I'll do that, thanks. I'm wondering, for example, if this is the expected behavior across all Sciex machines or not. It seems remarkable that this hasn't come up already.