Dilution factor per replicate waltteri hosia  2019-04-10
 

Hello the awesome Skyline team!

I am doing some impurity quantification, peptides. If I have made a calibration curve from pure target protein digest with ISTD concentration of, 400 µg/ml, and injected 10µl, and I have measured unknown series, but for few technical reason I had ISTD concentration of 100 µg/ml and injection volume of 3µl, then after merging files to same skyline document, how can I get quantification's of the unknown series? When trying to apply concentration multiplier in document grid, it applies automatically to all peptides with same sequence, i.e. also to calibration curve injections. Seems not be possible to apply it to unknown samples only?

 
 
Nick Shulman responded:  2019-04-10
You can specify the dilution factor for replicates in the Document Grid.
The "Dilution Factor" column does not appear by default, but you an add it to a view by using the "Views > Customize View" button on the Document Grid (in Skyline-Daily we have changed the name of "Views" to "Reports").

The Dilution Factor is a number that you set on unknown samples. When Skyline uses the calibration curve to compute the concentration of an unknown sample, Skyline multiplies the number by the Dilution Factor in order to arrive at the Calculated Concentration.

The "Concentration Multiplier" on peptides is something different. If the peptides in your stock solution are present in different quantities, you would use the Concentration Multiplier to make it so that each peptide's concentration is correct in your external standards. That is, the Analyte Concentration that you have specified on the external standard replicates times the Concentration Multiplier on the Peptide should be equal to the actual concentration of that peptide in that external standard replicate.
-- Nick