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Systematic evaluation of blood contamination in nanoparticle-based plasma proteomics

ABSTRACT: Circulating blood proteomics enables minimally invasive biomarker discovery. Nanoparticle-based circulating plasma proteomics studies have reported varying number of proteins (ca 2000-7000), but it’s unclear whether higher protein number is more informative. Here, we first develop OmniProt – a silica-nanoparticle workflow optimized through systematic evaluation of nanoparticle types and protein corona formation parameters. Next, we present an Astral spectral library for 10,109 protein groups. Using Astral with 60 sample-per-day throughput, OmniProt identifies ca 3000 to 6000 protein groups from human plasma. Notably, platelet/erythrocyte/coagulation-related contamination artificially elevates protein identifications and compromises quantification accuracy in nanoparticle-enriched samples. Through controlled contamination experiments, we identified biomarkers for platelet/erythrocyte/coagulation-related contaminations in nanoparticle-based plasma proteomics. We developed open-access software Baize for contamination assessment. We validated pipeline in 193 patients with CT-indistinct benign nodules or early lung cancers, flagging five contaminated samples. This study reveals contamination alters protein identification/quantification in nanoparticle-based plasma proteomics and presents Baize software to evaluate contamination.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Platelet, Blood Plasma, Blood Serum

SUBMITTER: 欢高

LAB HEAD: Tiannan Guo

PROVIDER: PXD068107 | Pride | 2025-12-12

REPOSITORIES: Pride

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			Action	DRS
	Astral_plasma_build_spectral_library_DIANN181report20241023-lib.tsv	Tabular
	CAF20240205gaohh_EV_24minDIA_30F_F1.raw	Raw
	CAF20240205gaohh_EV_24minDIA_30F_F10.raw	Raw
	CAF20240205gaohh_EV_24minDIA_30F_F11.raw	Raw
	CAF20240205gaohh_EV_24minDIA_30F_F12.raw	Raw

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Project description:Background: We aimed to investigate the effects of intravenous immune globulin (IVIG) and rituximab desensitization treatment on kidney transplant rate and blood gene expression profiles by microrarrays. Methods: We enrolled patients with PRA levels >50% and on the deceased-donor waiting list for >5 years. Patients received IVIG (2.0 g/kg) on day 0 and 30; and rituximab (375 mg/m2) on day 15. The antibodies with mean fluorescence intensity (MFI) values > 5,000 were reported to UNET as unacceptable antigens. The gene expression profiles of blood samples collected in PAXGene tube were studied by Affymetrix HuGene 1.0 ST expression arrays. Results: 40 of the 415 patients (10%) on the waiting list were eligible for desensitization treatment and 11 completed the treatment. While 15 of the remaining 29 patients (52%) received a transplant without therapy, only 2 of the 11 desensitized patients (18%) received transplant during a median follow-up of 217 days. While there were no statistically significant difference in demographics, desensitized patients had higher cPRA values (97% vs. 77%, p=0.0005) and more number of unacceptable antigens (39 vs. 10, p=0.0001). There was no significant change in the mean number of unacceptable antigens (39 ± 22 versus 39 ± 23) or reduction in the mean MFI values (11,333 ± 3,133 vs 11,289 ± 3,386). Analysis of genes chosen as significantly differentially expressed revealed downregulation of genes involved in B cells and immune system (CD79a, B and T lymphocyte associated transcript, B cell scaffold protein, CD22, CXCR5, fas apoptotic inhibitory protein). Gene set enrichment analysis using Pathogenesis Based Transcripts created by Edmonton Group demonstrated significant downregulation of B cell associated (p=0.04) and immunoglobulin transcripts (p=0.03). Conclusion: Although, desensitization with IVIG and rituximab decreases the expression of B cell and immunoglobulin associated transcripts, it was not successful in increasing kidney transplant rate or in decreasing the number of unacceptable antigens. Total of 28 arrays included in this study, which corresponding to 9 individuals with paired pre/post treatment samples and an additional 10 untreated control individuals. pair_analysis_normData.txt for paired analysis of pre/post treatment; ordinary_analysis_normData.txt for non-paired analysis include all samples, except for 38 Pax V0 and 56 Pax V1, which shows technicial bias and lowest array quality, hence removed from analysis.

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Systematic evaluation of blood contamination in nanoparticle-based plasma proteomics

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