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QuaSAR error jung165  2018-05-30 17:23
 

Hello Skyline team.

I have a one quick question about running AuDIT through QuaSAR (external tool) on Skyline.
However, I got LOD and LOQ calculation error during analysis and I am curious about solving this issue.

I already posted a question about this issue on this support board but, I got a reply that I have to ask it on support board on external tool.
So I posted another question on external tool of QuaSAR support board about 4 months ago but still could not get answer.
I also posted my question on GenePattern, developer of QuaSAR (I guess) and they said QuaSAR running through Skyline is not a tool they support.

Where should I ask about this problem??

Thank you for your help
 
 
Nick Shulman responded:  2018-05-30 20:38

Can you send us your Skyline document?

In Skyline, you can use the menu item:
File > Share > (complete)
to create a .zip file containing your Skyline document and supporting files including extracted chromatograms.

If that .zip file is less than 50MB, then you can attach it to this support request.
Otherwise, you can upload it here:
https://skyline.ms/files.url
 
jung165 responded:  2018-05-31 01:59

Thank you for reply

I attached zip file of my Skyline file.
If you run QuaSAR on tool tab, it would be possible to see that LOD calculation was halted.

I appreciate for your help
 
Nick Shulman responded:  2018-05-31 14:00

I am not very familiar with how QuaSAR is supposed to work, but when I run this through the debugger in R, it stops on a line of code where it thinks that you do not have any blanks.

The error message is telling you that you should disable the LOD calculation because you do not have any blanks. (indeed, all of the replicates in your Skyline document have the concentration specified as 3)

You can get QuaSAR to complete successfully if you uncheck "Plot each peptide" and "LOD/LOQ table" on the QuaSAR dialog that comes up when you run the QuaSAR external tool in Skyline. I do not know whether unchecking both of those checkboxes means that QuaSAR will not be doing the important part of its work.

I hope this helps!
 
jung165 responded:  2018-05-31 17:27

I appreciate for your kind reply.

Yes, it helps me well and I could get AuDIT result after unchecking those parameters.
After all, it brings me another question. Can I ask what does "blank" means for LOD/LOQ calculation?? If I analyzed my target proteins with spiking in SIS peptides, what should I use blank for calculating LOD/LOQ calculation??

Thank you for your help.
 
Nick Shulman responded:  2018-05-31 17:55
A blank would be a sample that you spiked your standards into, but which did not contain your analyte. (As opposed to a "double blank" which contains neither standards nor analyte)
 
jung165 responded:  2018-05-31 18:04
Thank you for your reply. It helps me a lot.

To make sure, If I spiked in SIS peptides, does blank would be the run only contains SIS peptides??
If so, does the number of replicate run of blank should be same as real analysis(sample with spike-in SIS peptide) or just single run of blank would help??

I appreciate for your help.