DIA analysis by IDSL.CSA
The Data-Independent Acquisition (DIA) peak processing approach presented in the DIA tab of the IDSL.CSA parameter spreadsheet for the IDSL.CSA package was designed to perform MS2 peak deconvolution on the MS1 peaks detected by IDSL.IPA. First, the fragmentation ion seeds are identified from the nearest raw MS2 chromatogram scan to the MS1 scan at the apex of MS1 chromatographic peaks. The MS1 and MS2 chromatograms are subjected to a two-step smoothing process, utilizing the LOESS function followed by cubic spline smoothing, aimed at increasing the number of points and reducing the jaggedness of the peaks. Pearson's correlation coefficients of the chromatograms of MS2 fragmentation peaks are calculated with respect to MS1 chromatogram peak. Finally, the mass of the MS1 chromatogram peaks as precursor value and correlating MS2 ions with the MS1 ion as fragmentation spectra are exported in a standard MSP format.
The IDSL.CSA/DIA chromatogram deconvolution approach is also capable of automatically detecting Sequential Window Acquisition of All Theoretical fragment ion spectra (SWATH-MS) DIA data types through the utilization of offset information present in the metadata of mass spectrometry files. In the absence of offset information, the IDSL.CSA/DIA defaults to the All-Ion Fragmentation (AIF) data analysis mode.
In Sequential Window Acquisition of All Theoretical fragment ion spectra (SWATH-MS) analyses, the offset range refers to the range of precursor ions selected for fragmentation in given MS2 SWATH-MS scans; therefore, multiple precursor ions may be included in a single MS2 SWATH-MS scan. Since only MS2 SWATH-MS scans with the same offset range are used for creating chromatograms, these MS2 SWATH-MS scans are distributed across many scans compared to MS1 scans of a chromatographic peak. Thus, to improve the quality of MS2 SWATH-MS chromatographic peaks, it is recommended to use higher MS2 smoothing windows. For example, when the smoothing window for MS1 was set to 25 MS1 scans, the smoothing window for MS2 scans should be set to 500 MS2 SWATH-MS scans.
Example 1: Chromatogram deconvolution for Pyridoxine in the MSV000090242 study
Example 2: Chromatogram deconvolution for Tryptophan in the MSV000090242 study
Hint: The spectral entropy (S) values are different between IDSL.CSA/DIA peak deconvolution and IDSL.FSA peak annotation because a 1% noise removal was applied in the spectra annotation step.
In All-Ion Fragmentation (AIF) analysis, all ions in the ion source of the mass spectrometry instruments are simultaneously fragmented, providing an unbiased and comprehensive view of the sample without the prior selection of precursor ions. However, the lack of precursor ion selection also limits the ability to accurately and efficiently search the reference libraries. To overcome this limitation with AIF DIA analysis, IDSL.CSA/DIA has been designed to establish connections between MS1 peaks of "intact" ions and MS2 peaks of fragmented ions, through mathematical correlations between the chromatography of MS1 and MS2 peaks. In essence, the IDSL.CSA/DIA workflow identifies instances where the chromatographic peaks of MS2 ions correlate with those of the MS1 ions, allowing for the corresponding MS1 peaks to be considered as precursor ions for the AIF DIA fragmentation spectra.
Example 1: Chromatogram deconvolution for Rutin in the MSV000087782 study
