Data-independent acquisition (DIA) is increasingly popular in mass spectrometry-based proteomics providing unbiased results and high data completeness. Adding an ion mobility dimension in diaPASEF on timsTOFpro mass spectrometers has proven effective for achieving deep proteome coverage, even with short liquid chromatography gradients commonly used in high throughput studies. However, for large sample cohorts, ensuring both mass stability (m/z) and ion mobility stability (1/K0) over time is crucial for consistent data quality.
We discovered that even minor changes in environmental air pressure during measurements systematically affect TIMS vacuum pressure, leading to shifted ion mobilities. By comparing experimental ion mobilities with historical weather data, we found that drifts observed over a few days can be attributed to typical day-to-day fluctuations in ground air pressure. Even small ion mobility drifts negatively impact the quality of quantitative results. Ions at the mobility boundaries of the diaPASEF precursor isolation windows exhibit drift-dependent abundance changes and increased missing values, which cannot be corrected by post-processing.
To address this issue, we applied an in-batch mobility autocalibration feature on a run-wise basis that robustly prevents weather-related ion mobility drifts.