Introduction
Data-independent acquisition-based SWATH-MS typically non-labelled protein samples are digested with trypsin and the resulting peptides are analyzed by liquid chromatography coupled to a tandem mass spectrometer operating in the so-called data-independent acquisition (DIA) mode[1]. In this mode, all ionized compounds of a given sample that fall within a specified mass range are fragmented in a systematic and unbiased fashion. The major advantage of SWATH-MS is that it supports quantitative analyses of peptides covering 1,000s of proteins with a high quantitative consistency and accuracy. It is ideally suited for projects that entail a large number of samples and that require accurate and reproducible quantification for the major fraction of the expressed proteome or peptidome in each sample.
Figure1 Principle of sequentially windowed data-independent acquisition in SWATH-MS[1].
In comparative de novo identification and quantification studies in human cell lysates, DIA identified up to 89% of the proteins detected in a comparable DDA experiment while providing reproducible quantification of over 85% of them. Comparison of DIA and DDA quantification as follows:
| Category | DIA | DDA |
|---|---|---|
| Definition | all ionized peptides of a given sample that fall within a specified mass range are fragmented in a systematic and unbiased fashion using rather large precursor isolation windows | single precursor ions are isolated, fragmented, and analyzed in an MS2 scan by the mass spectrometer. The precursor ions are chosen by the instrument on the basis of abundance |
| Repeatability of samples | better | poor |
| Application | large-scale quantification | small-scale quantification |
| Data analysis | complex | complex |
| Sensitivity and precision | higher | ordinary |
Bioinformatics Analysis Content
- Complete proteomics lab reports (including specific experimental procedures).
- Data processing
- Quality assessment of proteomic data
- Results of bioinformatics analysis
PCA analysis
Heat map analysis of all samples
Differential protein screening and analysis
Functional annotation of differential proteins
PPI analysis
How It Works
If you have any questions about our bioinformatics services, such as how we promote your research and build your reports, please contact us or online inquiry for more detailed information. Even if you don't have data yet, we can help you plan your research, provide expert experimental advice, and can schedule your data generation.
Q: How does a report looks like?
A: We will provide a publication-ready report for our customer. It documents the aims, workflow, methods, software tools used, results, conclusions, and references. If you want to an example reports please contact us.
Q: What does DIA bring to proteomics?
A: The promises of data-independent acquisition (DIA) strategies are a comprehensive and reproducible digital qualitative and quantitative record of the proteins present in a sample. All ionized peptides of a given sample and XIC information was collected for almost 100% of the primary + secondary ions. Compared to a data-dependent acquisition (DDA) experiments, DIA assay doubled the number of identified peptides and proteins per sample at half the coefficients of variation observed for DDA data (DIA = ∼8%; DDA = ∼16%).
Reference
- Ludwig, C., et al., Data-independent acquisition-based SWATH-MS for quantitative proteomics: a tutorial. Molecular Systems Biology, 2018. 14(8): p. e8126.
