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bioRxiv · 10.1101/2021.03.08.434385

High sensitivity dia-PASEF proteomics with DIA-NN and FragPipe

Demichev, V.·Yu, F.·Teo, G. C.·Szyrwiel, L.·Rosenberger, G.·Decker, J.·Kaspar-Schoenefeld, S.·Lilley, K. S.·Muelleder, M.·Nesvizhskii, A. I.·Ralser, M.
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Abstract

The dia-PASEF technology exploits ion mobility separation for high-sensitivity analysis of complex proteomes. Here, we demonstrate neural network-based processing of the ion mobility data, which we implement in the DIA-NN software suite. Using spectral libraries generated with the MSFragger-based FragPipe computational platform, the DIA-NN analysis of dia-PASEF raw data increases the proteomic depth by up to 69% compared to the originally published dia-PASEF workflow. For example, we quantify over 5200 proteins from 10ng of HeLa peptides separated with a 95-minute nanoflow gradient, and over 5000 proteins from 200ng using a 4.8-minute separation with an Evosep One system. In complex samples, featuring a mix of human and yeast lysates, the workflow detects over 11700 proteins in single runs acquired with a 100-minute nanoflow gradient, while demonstrating quantitative precision. Hence, the combination of FragPipe and DIA-NN provides a simple-to-use software platform for dia-PASEF data analysis, yielding significant gains in high-sensitivity proteomics.

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Demichev, V., Yu, F., Teo, G. C., Szyrwiel, L., Rosenberger, G., Decker, J., Kaspar-Schoenefeld, S., Lilley, K. S., Muelleder, M., Nesvizhskii, A. I., Ralser, M.. 2021-03-09. High sensitivity dia-PASEF proteomics with DIA-NN and FragPipe. https://doi.org/10.1101/2021.03.08.434385

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