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J Biomol Tech. 2010 September; 21(3 Suppl): S56.
PMCID: PMC2918053

Mining the Human Placenta Proteome > 5000 Proteins Deep Using CID/ETD on a Novel Ion Trap

S. Lemeer,1 A. Schneider,2 M. Lubeck,2 and B. Kuester1
1Technische Universität München and Center for Integrated Protein Sciences Munich, Germany;
2Bruker Daltonik GmbH, Bremen, Germany

Abstract

RP-112

To realise the potential of proteomics, it is important to be able to measure a proteome as completely as possible. Novel technology is providing important steps towards this goal and MS-based peptide sequencing in particular is showing great promise. To expand our knowledge about proteins expressed in human placenta - a primary human tissue of embryonic origin – we applied a combination of protein and peptide separation techniques in conjunction with CID and ETD on a novel ion trap o identify several thousand human proteins. Our data represents the most complete protein atlas available for primary human tissues to date. Post-delivery human placenta was obtained with written consent from a healthy female. Total protein extraction was performed using 6 M guanidine HCl, 1% v/v Triton X-100. Protein extracts were separated by 1D SDS gel electrophoresis and each lane cut into 18 regions. All samples were digested with trypsin. Peptides were separated by nanoscale RP-HPLC (Bruker Easy-nLC) coupled online to the Bruker amazon ion trap. MS/MS spectra were searched against the IPI database using Mascot and protein identifications were accepted at a false discovery rate of <5%. The improved scan speed of the instrument and a novel ion transfer line provided for a high duty cycle and sensitivity thus maximising the number of acquired high quality tandem mass spectra per unit time. Each of the elements of this multi-tier approach contributes to improved proteome coverage and collectively enabled the identification of several thousand proteins from human placenta, an improvement of a factor of at least two over traditional MudPit approaches.


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