Dataset Information

Comparative evaluation of different sample preparation workflows for reproducible, quantitative, and in-depth analysis of urine proteomics

ABSTRACT: The growing field of urinary proteomics has provided a promising opportunity to identify biomarkers useful for diagnosis and prognostication of a number of diseases. Urine is abundant and readily collected in a non-invasive manner which makes it eminently available for testing, however sample preparation for proteomics analysis remains one of the biggest challenges in this field. As newer technologies to analyze tandem mass spectrometry (MS) data develop, utility of urinary proteomics would be enhanced by better sample processing and separation workflows to generate fast, reproducible, and more in-depth proteomics data. In this study, we have evaluated the performance of four sample preparation methods: MStern, PreOmics In-StageTip (iST), Suspension-trapping (S-Trap), and conventional urea In-Solution trypsin hydrolysis for non-depleted urine samples. Data Dependent Acquisition (DDA) mode on QE-HF was used for single-shot label-free data acquisition. Our results demonstrate a high degree of reproducibility within each workflow. PreOmics iST yields the best digestion efficiency with lowest percentage of missed-cleavage peptides. S-trap workflow gave the greatest number of peptide and protein identifications. Using S-trap method, with 0.5mL urine sample as starting material, we identify ~1500 protein groups and ~17700 peptides from DDA analysis with a single injection. The continued refinement of sample preparation (presented here), LC separation methods and mass spectrometry data acquisition can allow the integration of information rich urine proteomic records across large cohorts with other ‘big data’ initiatives becoming more popular in medical research.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Hossein Fazelinia

LAB HEAD: Steven H. Seeholzer

PROVIDER: PXD016273 | Pride | 2020-03-06

REPOSITORIES: Pride

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Publications

Urine Proteomics: Evaluation of Different Sample Preparation Workflows for Quantitative, Reproducible, and Improved Depth of Analysis.

Ding Hua H Fazelinia Hossein H Spruce Lynn A LA Weiss Dana A DA Zderic Stephen A SA Seeholzer Steven H SH

Journal of proteome research 20200310 4

The growing field of urinary proteomics shows promise to expand the number of biomarkers for the diagnosis and prognosis of a number of human diseases. With the rapid developments in mass spectrometry methods for proteome quantification, there exists an opportunity for improved sample processing and separation workflows to make important contributions to urine proteomic analyses. Here we evaluate the performance of four sample preparation methods: MStern, PreOmics in-StageTip (iST), suspension-t ...[more]

PMID: 32129078

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Project description:Total pancreatectomy with islet cell autotransplantation (TPIAT) is a surgical treatment modality used for the management of painful chronic pancreatitis when medical and endoscopic therapies fail. Total pancreatectomy (TP) results in a number of clinical implications. However, the glucose-responsive insulin secretion can be preserved by combining TP with islet cell autotransplant (IAT). Despite the known clinical implications of TPIAT, the systemic molecular effects of TPIAT remains poorly investigated. Therefore, we performed the first hypothesis-generating study of the urinary proteome before and after TPIAT. The RAW- and processed data from the analysis can be found in this repository. Twenty-two chronic pancreatitis patients eligible for TPIAT were prospectively enrolled in the study at the University of Minnesota Medical Center between September 2011 and February 2014. Urine was collected within the week pre-TPIAT and 12-18 months post-TPIAT, resulting in 44 paired samples. The paired experimental design allowed us to subtract interpersonal differences. The unique set of urine samples were prepared for bottom-up label-free quantitative proteomics using the in-house developed ‘MStern’ protocol for the 96-well plate compatible processing of the urine samples. The samples were analyzed by LC-MS/MS and 2477 proteins were identified (FDR<0.01%). Five samples were found to have less than 30% of the cumulated list of proteins (<743 proteins) and so these samples and their respective matching sample were thus removed from the analysis. This resulted in a complete dataset with 17 matched pre- and post-TPIAT sample pairs. The final list of recommended included pairs for future analysis are indicated in the PX_TPIAT_Metadata.csv file. Note that extended clinical information only was available for the included samples. Furthermore, the information is only listed for the pre-TPIAT sample but applies to both the pre- and post-TPIAT participant sample. The data allows for studying the systemic molecular impact of the TPIAT procedure.

Dataset Information

Comparative evaluation of different sample preparation workflows for reproducible, quantitative, and in-depth analysis of urine proteomics

Publications

Urine Proteomics: Evaluation of Different Sample Preparation Workflows for Quantitative, Reproducible, and Improved Depth of Analysis.

Similar Datasets

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