Incorrect Peak Area Sum when Using Sum Multiple Ions Feature in MultiQuant Software 3.0.1


日付: 03/22/2018
カテゴリー: MultiQuant Software

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For research use only. Not for use in diagnostic procedures.


Answer

When analyzing MRMHR data, selected fragment ions of a precursor ion can be summed using the Sum Multiple Ions feature in MultiQuant™ 3.0.1 software with Hotfix 2 (see example below):
sum multiple ions
In the above example, the two fragment ions, 626.4 and 608.2, were summed using the Sum Multiple Ions algorithm by right clicking on the row.

sum multiple ion feature
The results of running Sum Multiple Ions were added to the results table (see line above Res 3). The summing added an extra column named "Start-Stop-2". Next  the peak areas were compared to those calculated using Excel.



When Excel was used to calculate the sum of the XIC peak areas for the two fragment ions, the resulting sum did not match the sum of the two peak areas that were calculated using the command Sum Multiple Ions. The reasons are explained below:

1) The peaks areas from different retention times were compared, which creates a problem calculating the peak area value correctly. The retention time was manually selected for the second compound (Res 2) as 4.8 min, but the RT of the third compound (Res 3) was slightly different, because the RT of Res 1 was different to begin with. Therefore, the sum of the peak areas for Res 1 and 2 can only be correct if they have the same RT.

2) This algorithm used is called Sum Multiple Ions not Sum Multiple Peaks. In short, the instrument response (intensity) from each time point of each individual ion transition is summed, and a completely new trace is created with those summed instrument responses. Then the software finds and integrates the peaks in this new trace using the user-defined or default integration parameters. The software does not add peak areas together from different peaks. When comparing results, all peaks must be from same retention time. Based on the nature of the summing logic, even peaks that are from the same retention time, the new trace might have different signal-to-noise and the peak start/stop might be different for the individual mass transition. Therefore, the peak integration could be different, resulting in peak areas that are not identical to the peak area sum from the individual peaks.