AlphaDIA enables DIA transfer learning for feature-free proteomics.

The scale of data generated for mass-spectrometry-based proteomics and modern acquisition strategies poses a challenge to bioinformatic analysis. Search engines need to make optimal use of the data for biological discoveries while remaining statistically rigorous, transparent and performant. Here we present alphaDIA, a modular open-source search framework for data-independent acquisition (DIA) proteomics. We developed a feature-free identification algorithm that performs machine learning directly on the raw signal and is particularly suited for detecting patterns in data produced by time-of-flight instruments. Benchmarking demonstrates competitive identification and quantification performance. While the method supports empirical spectral libraries, we propose a search strategy named DIA transfer learning that uses fully predicted libraries. This entails continuously optimizing a deep neural network for predicting machine-specific and experiment-specific properties, enabling the generic DIA analysis of any post-translational modification. AlphaDIA provides a high performance and accessible framework running locally or in the cloud, opening DIA analysis to the community.