Products/Tecan

Smile! Protein and peptide sample preparation automated

Application for high throughput proteomic sample preparation for LC-MS analyses using iST and SP3

The challenge:

How do you automate sample preparation of very dirty samples or where harsh denaturation reagents are needed? Typically these samples cause issues with proteomic sample preparation as the reagents can adversely impact digestion and subsequent LC-MS analyses.

The solution:

Use the combined advantages of SP3 and iST! Together, PreOmics and Tecan® have developed an automated LC-MS sample preparation workflow, combining the ability to wash and digest samples when bound to the SP3 beads and then do the clean-up step. This application meets the demands of efficient, robust proteomics sample preparation for difficult samples for LC-MS analysis with the bonus that an automated workflow helps labs decrease the hands-on time and produce high quality samples for LC-MS analysis!

“Tecan” is a registered Trademark of a third party

Relax! SP3-iST compatible workflows

1.
Adding samples

Samples should be liquid, homogenates or a soft pellet.

2.
SP3-iST kit set up

Prepare the SP3 beads and the DIGEST and add all reagents to the Tecan instruments.

3.
Sample preparation

Wash, lyse and digest samples while bound to SP3 beads on the Tecan Freedom EVO®.

4.
Sample clean-up

Samples are automatically transferred to the clean-up plate and processed on the Tecan Resolvex® A200.

5.
MS and data analysis

Analyse samples by
LC-MS and process the data.

For technical data and performance statistics see the SP3-iST application note and poster.

SP3-iST & iST-PSI | Automation of the SP3-iST PreOmics workflow on the Freedom EVO® 100 and the Resolvex® A200 | Tecan

SP3-iST | High-throughput automation of LC-MS sample preparation: The SP3-iST workflow on the Freedom EVO® 100 and the Resolvex® A200

Results

Table 1: Comparison of nanoLC-MS results for the SP3-iST automated and manual protocols.

Additional data can be found in the Tecan® SP3-iST application note. The numbers of PSMs and IDs for both peptides and proteins were very consistent for all samples, regardless of protein input, as well as between the manual and the automated protocols. Approximately 27,370 peptides and 2,960 proteins were identified in all samples, in agreement with the expected performance of the nanoLC-MS measurements conducted. This demonstrates the high efficiency, quality and robustness of the automated SP3-iST workflow for the entire range of tested input amounts (15-50 μg).