Priority Research Paper Volume 15, Issue 20 pp 10821—10855

Figure 1. Schematic of workflow. Ovaries were harvested from reproductively young (6–12 weeks) and reproductively old (10–12 months) mice. One ovary per mouse was kept native and the contralateral ovary was enriched for the ECM by treatment with 0.1% SDS for 12.5 h. Native and ECM-enriched ovaries were characterized by H&E, Hoechst, and PSR staining of tissue sections, as well as measurement of DNA fragment size on an agarose gel. Tissue was homogenized and proteins were extracted, digested using S-Trap, and desalted by HLB C₁₈ cartridges. Peptides were analyzed on an Orbitrap Exploris 480 by Data-Independent Acquisition (DIA)-MS to quantify dynamic protein changes across reproductive aging. Raw data were analyzed using Spectronaut and significantly altered proteins were subjected to pathway analysis using ConsensusPathDB-mouse.