Dear Skyline-Team,
I was trying to get an idea of how reliable the quantification of peptides/proteins using different DIA Methods would be.
As a first and major read-out I tried to look at the points across peak value. If i understand this parameter correctly it should give me the number of points leading to the determined Intensity / area of each transition / precursor.
As a first start I modified the MSStats Input report by adding the PAP and filtering for "is not blank".
Using the Pivot Editor I focused on proteins with a max Detected Q value below 0.01 (see attached pdf showing screenshots).
Further pivot editing led to Mean Points across peaks per Protein for each replicate in each of the thee different conditions.
As a first and quick step I filtered for each replicate within each conditions and calculated the Average of the Mean Points across Peak values (shown in the table).
Although the values show the expected trend (higher values with shorter cycle time due lower max IT). The average values between 14 -16 seem to be a little high.
When going through the data I noticed that the integration boundaries for some peptides are nicely selected (e.g. AIGPHDVLATLLNNLK) but on the other hand for some peptides the integration boundaries seem to be too broad/ unspecific selected (e.g. LQTSSVLVSGLR).
Is there a way to specify global settings for peak integrations boundaries, e.g. more conservative or less conservative / more sensitive? Or does one have to go through all peptides manually and adjust the boundaries?
Best regards
Matthias