Heavy peptide as global or surrogate standard

Heavy peptide as global or surrogate standard waltteri hosia  2019-09-20

Hi, Skyline keeps coming better with every update, thank you for keep improving it.

Question. Is there a way to set the heavy peptide as a global or surrogate standard? I have an quant experiment with few proteins and I only have one heavy peptide per protein. I see though that sensitivity would be better with other peptide than the one that I have the heavy for. The heavy is in as one fixed concentration.

Nick Shulman responded:  2019-09-20
Yes, you can do this, but you will have to set the Standard Type of each of these heavy peptides to "Surrogate Standard" and then set the "Normalization Method" for all the other peptides to the appropriate "Ratio to Xxxx".

The "Fill Down" right click menu item in the Document Grid will probably be helpful for this.
-- Nick
waltteri hosia responded:  2019-09-20
Hi, Thanks for quick response. I couldn't quite do this. When you have light and heavy they are under the one same sequence and skyline doesn't show the "set standard type" option for light/heavy, only for whole peptide -including both. But I could delete the light and set the heavy that's left as global standard. Works! (Saved then Skyline document with new name XXX_heavy as global) BR, Waltteri
Nick Shulman responded:  2019-09-20
For "Set Standard Type", you should mark the entire peptide as a Surrogate Standard.

When a surrogate standard has both a light and heavy form, there will be multiple options in the Normalization Method column for choosing whether you want to use the entire peptide, or just the light or heavy forms. That is, you should see:
Ratio to surrogate PEPTIDEK
Ratio to surrogate PEPTIDEK (light)
Ratio to surrogate PEPTIDEK (heavy)

-- Nick
waltteri hosia responded:  2019-09-24

Delayed thank you! Very helpful.