Abstract
Large-scale, unbiased proteomics studies are constrained by the complexity of the plasma proteome. Here we report a highly parallel protein quantitation platform integrating nanoparticle (NP) protein coronas with liquid chromatography-mass spectrometry for efficient proteomic profiling. A protein corona is a protein layer adsorbed onto NPs upon contact with biofluids. Varying the physicochemical properties of engineered NPs translates to distinct protein corona patterns enabling differential and reproducible interrogation of biological samples, including deep sampling of the plasma proteome. Spike experiments confirm a linear signal response. The median coefficient of variation was 22%. We screened 43 NPs and selected a panel of 5, which detect more than 2,000 proteins from 141 plasma samples using a 96-well automated workflow in a pilot non-small cell lung cancer classification study. Our streamlined workflow combines depth of coverage and throughput with precise quantification based on unique interactions between proteins and NPs engineered for deep and scalable quantitative proteomic studies.
Highlights
Background robustness testInterference substances were obtained from Sun Diagnostics
They may not be suitable for high-throughput translational proteomic analysis due to the necessity of repeated centrifugation or membrane filtration to separate the corona from free plasma proteins, and to wash away loosely attached proteins
SPIONs can be robustly modified with different surface chemistries, which may facilitate the generation of distinct corona patterns for broader interrogation of the proteome (Supplementary Fig. 2)
Summary
Interference substances were obtained from Sun Diagnostics. Hemolysate: Red blood cell hemolysate derived from human. A pooled plasma was spiked at different concentrations Lipid: High (1000 mg/dL), Low (100 mg/dL), and Control (buffer only). Hemolysate: High (1000 mg/dL), Low(100 mg/dL), and Control (buffer only). Statistical analysis and visualization were performed using R (v3.5.2) with appropriate packages[74]. Experiments were conducted in assay replicates (n = 3) unless noted differently. NSCLC data were acquired for biological replicates (see above). Mass spectrometry raw data and functional protein annotation references are available through PRIDE75 and Perseus[76], respectively
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