Comparative Evaluation of DDA and DIA Based Proteomic Workflows in Beryllium Related Lung Disease

Weise, D. O.; Gupta, K.; Griffin, T. J.; Jagtap, P. D.; Mroz, M. M.; Wagner, R.; Macaluso, J. D.; Mehta, S.; Maier, L. A.; Li, L.; Vestal, B. E.; Bhargava, M.

doi:10.64898/2026.06.04.730108

bioRxiv · 10.64898/2026.06.04.730108

Comparative Evaluation of DDA and DIA Based Proteomic Workflows in Beryllium Related Lung Disease

Weise, D. O.·Gupta, K.·Griffin, T. J.·Jagtap, P. D.·Mroz, M. M.·Wagner, R.·Macaluso, J. D.·Mehta, S.·Maier, L. A.·Li, L.·Vestal, B. E.·Bhargava, M.

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Abstract

We compared traditional data-dependent acquisition mass spectrometry (DDA-MS) with the increasingly adopted data-independent acquisition (DIA-MS) to evaluate their relative utility for large-scale quantitative biofluid proteomics of lung compartments, specifically paired bronchoalveolar lavage (BAL) cells and bronchoalveolar lavage fluid (BALF). Using beryllium-related granulomatous lung disease as a focused model, we analyzed BALF and BAL cells from beryllium-sensitized (BeS) individuals using both acquisition strategies to assess proteome depth, quantitative completeness, and analytical robustness. In BAL cells, 5,640 proteins were identified by DDA-MS and 5,227 by DIA-MS; however, DIA-MS yielded markedly improved quantitative completeness, with 5,178 proteins ([~]99%) quantified across all samples compared with 3,539 ([~]63%) quantified by DDA-MS. While 3,397 proteins were quantified by both methods, DIA-MS uniquely quantified 1,781 lower-abundance proteins. Proteins identified by both DIA and DDA-MS approaches revealed pathways associated with granulomatous inflammation, including Toll-like receptor, clathrin-mediated endocytosis, sirtuin, and C-type lectin receptor signaling, whereas DIA-MS resolved additional pathways, such as the complement cascade, coagulation system, and JAK/IL-6-type cytokine signaling. In BALF, although more proteins were identified by DDA-MS than by DIA-MS (2,069 vs 1,742), DIA-MS achieved greater quantitative completeness, with 1,695 proteins quantified across all samples compared with 1,050 using DDA-MS, underscoring its suitability for biomarker-oriented analyses in lung fluid compartments. Together, these results support DIA-MS as a robust and sensitive platform for quantitative lung proteomics and discovery of disease-relevant protein signatures.

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Weise, D. O., Gupta, K., Griffin, T. J., Jagtap, P. D., Mroz, M. M., Wagner, R., Macaluso, J. D., Mehta, S., Maier, L. A., Li, L., Vestal, B. E., Bhargava, M.. 2026-06-09. Comparative Evaluation of DDA and DIA Based Proteomic Workflows in Beryllium Related Lung Disease. https://doi.org/10.64898/2026.06.04.730108

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