Postby Zhang » Thu Jan 16, 2014 9:00 am
The reason why they load 300ng is further increase on-column loading will decrease the performance, not like the Orbi ones.
However, in Orbi experiments, with the 60 ms max-injection-time they almost never let the instrument to accumulate enough ions into collision cell. Besides, they used so wide a scan range in MS1, 300-1750. I mean seriously, how many peptides can be identified under 400.0 Th or over 1400Th? At the same time, they used two lock masses in every scan, MS1 and MS2, which significantly slowed down the data acquisition.
In my opinion, higher injection but lower identification means, when complexity goes up DIA will fail. However, DIA was designed for high complexity in handles multiplexed MS/MS. And the number game is also questionable, because what you search from DDA is the complete proteome sequence database, but for DIA it's only an MS/MS database.
I agree the one hour yeast is as well playing number games, and the different organism could not be directly compared, so is DIA vs DDA, and so is TOF vs Orbi. I was arguing that the statement in this DIA paper is misleading, trying to convince people that DIA is a superior method that beats DDA from every aspect. Shame on the reviewers, they agreed!