Age-Associated Plasma Protein Signatures and Proteomic Age Estimation by Data-Independent Acquisition Mass Spectrometry in a Russian Cross-Sectional Cohort.

Chronological age can be estimated from molecular profiles; however, predictive accuracy alone does not establish a biologically validated ageing clock. We therefore evaluated whether plasma protein intensities measured by data-independent acquisition mass spectrometry (DIA-MS) support chronological-age estimation in a cross-sectional cohort from Moscow, Russia, while explicitly examining acquisition-order and MS-batch confounding. Plasma samples from 350 adult participants aged 19.8 to 99.1 years were analyzed as single-shot, single-injection DIA-MS acquisitions on a Q Exactive HFX platform. The final matrix comprised 222 protein groups and was generated with DIA-NN version 1.8.1 using a two-pass predicted-library-assisted workflow, cross-run precursor-level normalization, normalized MaxLFQ-like protein-group quantification, and 1% precursor and protein-group false-discovery-rate thresholds. Descriptive associations were assessed using Spearman rank correlation with Bonferroni correction. Feature selection and missing-value imputation were confined to each training subset. Generalization was assessed primarily by holding out complete MS batches, with repeated, nested, and random five-fold analyses retained as internal comparators. Of the 222 proteins, 148 were associated with chronological age at the nominal threshold