Ghaffari, B. | Search bioRxiv

bioRxiv2026

Deep nanoparticle protein corona plasma proteomics resolves a stage-specific peripheral signature of Alzheimer's disease

Structured AbstractO_ST_ABSINTRODUCTIONC_ST_ABSAlzheimers disease (AD) progresses over decades, yet plasma biomarkers that resolve disease stage rather than simply detect disease remain scarce. This distinction is clinically consequential because effective AD intervention depends on identifying patients before disease biology has progressed beyond a therapeutically responsive stage. METHODSWe used small-molecule-modulated protein corona proteomics to profile plasma from 90 individuals in the Australian Imaging, Biomarker and Lifestyle cohort, stratified by Centiloid (CL) A{beta}-amyloid burden (30 amyloid- negative, CL < 15; 30 moderate-to-high, CL 26 to 100; 30 very high, CL > 100). We quantified 3,176 proteins and applied differential abundance and actual causality analyses to identify stage-specific and candidate causal proteins. RESULTSDifferential protein abundance was exclusively captured during the moderate-to-high AD transition, revealing a discrete proteomic "switch." The switch was marked by accumulation of the autophagy receptor CALCOCO1, together with coordinated depletion of the S100A8/S100A9 calprotectin complex and core erythroid-cytoskeletal network structural markers (e.g., SPTA1, SPTB, ANK1). Adhesion G protein-coupled receptor G6 (ADGRG6) showed a significant moderate positive monotonic association with absolute CL burden, providing a proportional molecular anchor for cumulative disease burden. Actual causality analysis identified COL6A2, FOXRED2, P3H1, PRR4, and GOLGA5 as candidate upstream drivers linking matrix remodeling, Golgi trafficking, and collagen processing to AD progression. DISCUSSIONThese findings suggest a candidate blood-accessible framework for staging AD by active disease biology, which, if replicated in independent cohorts, may have implications for therapeutic selection and mechanism-guided clinical trials. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=92 SRC="FIGDIR/small/740710v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1ead718org.highwire.dtl.DTLVardef@cfa523org.highwire.dtl.DTLVardef@62a367org.highwire.dtl.DTLVardef@1d5c9a5_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

Recalibrating Nanoparticle Protein Corona Analysis for Accurate Biological Identity and Soluble Plasma Proteome Profiling

The nanoparticle (NP) protein corona is considered the biological identity that determines NP fate, safety, targeting, and therapeutic effectiveness in biofluids. Nonetheless, standard corona isolation workflows assume the recovered protein signature originates primarily from plasma proteins adsorbed directly onto the NP surface, while largely overlooking co-isolation of endogenous nanoscale biological structures such as extracellular vesicles (EVs). This oversight can distort the apparent "biological identity" of the NP. Here, we show that EVs are a major hidden contributor to the perceived protein corona composition in human plasma. Using highly monodispersed polystyrene NPs (50-1000 nm) and superparamagnetic beads, we compared corona formation in standard human plasma and plasma depleted of an EV- enriched sedimentable fraction by ultracentrifugation at 100,000 x g for 2 h, with the recovered vesicles subsequently characterized by MACSPlex immunoaffinity analysis. Mass spectrometry revealed that EV depletion reduced the number of proteins identified on polystyrene NPs by 60-75% and on magnetic beads by 45-50%, demonstrating a substantial fraction of the conventionally assigned corona proteome arises from EV- associated carryover. EV depletion also restructured the apparent abundance hierarchy, increasing the relative prominence of soluble plasma proteins such as albumin and shifting dominant signals away from intracellular cytoskeletal component proteins that are characteristic of EV carryover towards genuine soluble plasma proteins and complement factors. These results highlight that standard corona workflows can inadvertently co-isolate a vast array of EV-associated material and thereby yield inaccurate assignments of protein origin. Distinguishing proteins adsorbed from the soluble phase from EV-surface and intravesicular material is essential for accurate interpretation of NP-biofluid interactions, biomarker discovery, and therapeutic targeting because molecular compartment determines both analytical meaning and drug accessibility. Significance StatementThe nanoparticle (NP) "protein corona" defines how engineered nanomaterials interact with living systems, influencing therapeutic safety, efficacy, and diagnostic utility. Conventionally, corona isolation workflows assume that recovered proteins were adsorbed directly from the fluid phase. This study reveals a major, previously overlooked source of analytical distortion: standard separation techniques routinely co-isolate endogenous extracellular vesicles (EVs), drastically distorting the perceived biological identity of NPs. Depletion of an EV-enriched sedimentable fraction by ultracentrifugation reduced identified corona proteins by up to 75% and restructures the apparent proteomic hierarchy. Distinguishing true soluble adsorbates from vesicular carryover is essential for accurately predicting NP behavior in vivo and prevents false positives in nano- diagnostics, establishing a critical new standard for high-fidelity biomarker discovery and nanomedicine.

bioengineering↗