1. Accelerating Research in Life Sciences
PREMIER Biosoft
Accelerating Research in Life Sciences

2. A High Throughput Lipid Characterization Tool using MS Data

3. Major Features Database containing 40,298 lipid structures.
5211 set masses (transitions) of PIS/NLS experiments for fatty acid chains, and head groups of lipid species from different classes and sub-classes.
Identify target lipids based on exact mass database search, and the target headgroup, or fatty acid chain corresponding to the set masses of the PIS/NLS experiments.
Isotope peak correction to facilitate accurate quantitation of lipids.

4. Major Features
Align lipids across biological samples based on short name e.g., PC(28:2); name without double bond positions e.g., PC(18:1/18:1) instead of PC(18:1(9Z)/18:1(9Z)).
Calculate normalized response of analytes w.r.t. the response of the internal standards that belong to the same class/subclass.
Heatmap, Concentration X Composition reports of lipid classes, and fatty acid chains.
Additional lipid ontology based filters, e.g., only even/odd numbers of double bonds, and carbon (C) atoms in the fatty acid chains of target lipids.

5. Create a New Project 1. File > New > Project
2. Enter a project name
3. Click Create

6. Import Data from Native as well as Standard Data Files
1. File > Open > Peaklist Data > Waters Data File > QqQ (.raw)

7. Shortcut for Importing “Waters” Data
1. Select Waters > QqQ (.raw)
2. Select the file(s)
3. Click Open

8. Enter PIS/NLS Target Masses for Each Function Within a File
2. To average MS2 level data select “Specify Range” option
3. Click OK
1. Click “All Scans of All Profiles” to select all the scans in one go

9. Enter PIS/NLS Target Masses for Each Function Within a File
1. Click Load Scan

10. Enter PIS/NLS Target Masses for Each Function Within a File

11. Model Experiment Design for Targeted Lipid Identification
2. Analyze > Lipid Quantitation > Model Experiment Design
1. Select Lipid Quantitation Node

12. Model Experiment Design for Targeted Lipid Identification
1. Select “Only Average Profiles” for experiment design
2. Click OK

13. Model Experiment Design
1. Specify experiment name
2. Specify sample name
4. Select the corresponding Events/Functions
3. Select the samples
6. Click to transfer all the information into selected profiles table
5. Polarity and Target Mass and Scan Type information is read automatically from the file

14. Model Experiment Design
Information transferred to selected profiles table
2. Click OK
1. Click “Map Target Mass Database” option to map the
target masses with SimLipid's target fragment database

15. Target Mass Mapping – Review the Mapped Fragments
1. Select this view to show only the
Events /files with unique target masses
within a specified tolerance range
2. Select target mass

16. Target Mass Mapping – Review the Mapped Fragments
Search results obtained for Target Mass =
Click OK
Information selected for this target mass is applied to all other instances of this target mass available in the “Show All” view.

17. Specify Peak Correction Parameters and Click “OK” to Perform Search
1. Select to enable Peak Correction
3. Click OK
2. Click Peak Correction Technique

18. Specify Peak Correction Parameters and Click “OK” to Perform Search

19. Data Pre-processing

20. Check Search Status and Load Results
HTP search status- Pending

21. Check Search Status and Load Results
HTP search status- Completed

22. Check Search Status and Load Results
Load HTP Results

23. Check Search Status and Load Results

24. Result View of PIS/NLS_253.2168_Pos
Notice the hierarchy of experiment, sample and and replicates

25. Result View at PISNLS_184.1_Pos
Note that lipids from PC and SM classes are only reported

26. Mark Internal Standard
1. Select a lipid to be marked as internal standard
2. Select Mark/Unmark/Edit Internal Standard button
4. Click Mark
3. Specify the amount of Internal Standard
Please Note: Multiple internal standards can be marked for a lipid class

27. Align Scans in a Sample
Analyze > Lipid Quantitation > Align Scans in Sample
Select the experiment name from the dropdown list
Select the PIS/NLS/Full Scan nodes to be aligned for a sample

28. Align Scans in a Sample

29. Scans Aligned at Sample Level
Note here lipid PC(44:0) identified in both scans with Target Mass = (Neg mode) and Target Mass
= (Pos mode) gets aligned, you can SUM or Average the intensities from the aligned scans

30. Align Samples in an Experiment
Use shortcut button to align samples
Analyze > Lipid Quantitation > Align Samples
1. Select the Experiment Name
2. Select the Samples to be aligned
3. Select the Alignment Options
4. Click OK

31. Align Samples in an Experiment

32. Generate Comp. x Conc. Report
2. Select the fragment type and lipid abundance option for generating Comp. X Conc. report
3. Select “Heat Map” to generate heatmap
1. Click on “Comp. X Conc. Report” button
4. Select file type
5. Click OK

33. Generate Comp. x Conc. Report

34. Generate Comp. x Conc. Report Output

35. Lipids Aligned at Sample Level
Lipids aligned from both the samples, Serum 10 and Serum 33

36. Quantitation at Experiment Level
2. Select Total Abundance
1. Click Quantitation
3. Click OK

37. Quantified Lipids at Experiment Level