Abstract
Mass spectrometric plasma analysis for biomarker discovery has become an exploratory focus in proteomic research: the challenges of analyzing plasma samples by mass spectrometry have become apparent not only since the human proteome organization (HUPO) has put much emphasis on the human plasma proteome. This work demonstrates fundamental proteomic research to reveal sensitivity and quantification capabilities of our Peptidomics technologies by detecting distinct changes in plasma peptide composition in samples after challenging healthy volunteers with orally administered glucose. Differential Peptide Display (DPD) is a technique for peptidomics studies to compare peptides from distinct biological samples. Mass spectrometry (MS) is used as a qualitative and quantitative analysis tool without previous trypsin digestion or labeling of the samples. Circulating peptides ( < 15 kDa) were extracted from 1.3 mL plasma samples and the extracts separated by liquid chromatography into 96 fractions. Each fraction was subjected to MALDI MS, and mass spectra of all fractions were combined resulting in a 2D-display of > 2,000 peptides from each sample. Endogenous peptides that responded to oral glucose challenge were detected by DPD of pre-and postchallenge plasma samples from 16 healthy volunteers and subsequently identified by nESI-qTOF MS. Two of the 15 MS peaks that were significantly modulated by glucose challenge were subsequently identified as insulin and C-peptide. These results were validated by using immunoassays for insulin and C-peptide. This paper serves as a proof of principle for proteomic biomarker discovery down to the pM concentration range by using small amounts of human plasma.
Keywords: Mass spectrometry, blood, peptidomics, differential display, glucose challenge
Combinatorial Chemistry & High Throughput Screening
Title: Prerequisites for Peptidomic Analysis of Blood Samples: II. Analysis of Human Plasma after Oral Glucose Challenge - A Proof of Concept
Volume: 8 Issue: 8
Author(s): Harald Tammen, Rudiger Hess, Imke Schulte, Markus Kellmann, Annette Appel, Petra Budde, Hans-Dieter Zucht and Peter Schulz-Knappe
Affiliation:
Keywords: Mass spectrometry, blood, peptidomics, differential display, glucose challenge
Abstract: Mass spectrometric plasma analysis for biomarker discovery has become an exploratory focus in proteomic research: the challenges of analyzing plasma samples by mass spectrometry have become apparent not only since the human proteome organization (HUPO) has put much emphasis on the human plasma proteome. This work demonstrates fundamental proteomic research to reveal sensitivity and quantification capabilities of our Peptidomics technologies by detecting distinct changes in plasma peptide composition in samples after challenging healthy volunteers with orally administered glucose. Differential Peptide Display (DPD) is a technique for peptidomics studies to compare peptides from distinct biological samples. Mass spectrometry (MS) is used as a qualitative and quantitative analysis tool without previous trypsin digestion or labeling of the samples. Circulating peptides ( < 15 kDa) were extracted from 1.3 mL plasma samples and the extracts separated by liquid chromatography into 96 fractions. Each fraction was subjected to MALDI MS, and mass spectra of all fractions were combined resulting in a 2D-display of > 2,000 peptides from each sample. Endogenous peptides that responded to oral glucose challenge were detected by DPD of pre-and postchallenge plasma samples from 16 healthy volunteers and subsequently identified by nESI-qTOF MS. Two of the 15 MS peaks that were significantly modulated by glucose challenge were subsequently identified as insulin and C-peptide. These results were validated by using immunoassays for insulin and C-peptide. This paper serves as a proof of principle for proteomic biomarker discovery down to the pM concentration range by using small amounts of human plasma.
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Tammen Harald, Hess Rudiger, Schulte Imke, Kellmann Markus, Appel Annette, Budde Petra, Zucht Hans-Dieter and Schulz-Knappe Peter, Prerequisites for Peptidomic Analysis of Blood Samples: II. Analysis of Human Plasma after Oral Glucose Challenge - A Proof of Concept, Combinatorial Chemistry & High Throughput Screening 2005; 8 (8) . https://dx.doi.org/10.2174/138620705774962427
DOI https://dx.doi.org/10.2174/138620705774962427 |
Print ISSN 1386-2073 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5402 |
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