LanthanideMetal–Organic Framework Flowersfor Proteome Profiling and Biomarker Identification in UltratraceBiofluid Samples

Identifying effective biomarkers has long been a persistent need for early diagnosis and targeted therapy of disease. While mass spectrometry-based label-free proteomics with trace cell has been demonstrated, deep proteomics with ultratrace human biofluid remains challenging due to low protein concentration, extremely limited patient sample volume, and substantial protein contact losses during preprocessing. Herein, we proposed and validated lanthanide metal–organic framework flowers (MOF-flowers), as effective materials, to trap and enrich protein in biofluid jointly through cation−π interaction and O–Ln coordination. We further developed a <b>M</b>OF-flower <b>a</b>ssisted <b>s</b>implified and <b>s</b>ingle-pot <b>S</b>ample <b>P</b>reparation (Mass-SP) workflow that incorporates protein capture, digest, and peptide elute into one single PCR tube to maximally avoid adsorptive sample loss. We adopted Mass-SP to decipher aqueous humor (AH) proteome signatures from cataract and retinal vein occlusion (RVO) patients and quantified ∼3900 proteins in merely 1 μL of AH. Combined with machine learning, we further identified PFKL as a prioritization biomarker for RVO disease with the areas under the curves of 0.95 ± 0.04. Mass-SP presents a strategy to identify de novo biomarkers and explore potential therapeutic targets with extremely limited clinical human body fluid resources.