Applying multiple proteases to direct digestion of hundred‐scale cell samples for proteome analysis. (5th July 2015)

• Record Type: Journal Article
• Authors: Chen, Qi; Yan, Guoquan; Zhang, Xiangmin
• Is Part Of: Rapid communications in mass spectrometry. Volume 29:Number 15(2015)
• Page Start: 1389
• Page End: 1394
• Publication Date: 2015-07-05
• Subjects: Mass spectrometry -- Periodicals; 543.65
• DOI: 10.1002/rcm.7230
• Languages: English
• ISSNs: 0951-4198

Abstract

<abstract abstract-type="main"> <title> <x xml:space="preserve">Abstract</x> </title> <sec id="rcm7230-sec-0001" sec-type="section"> <title>Rationale</title> <p>Analyzing the proteome on the scale of only several hundred cells with mass spectrometry has great significance for applications with limited sample amounts. We applied multiple proteases to the direct digestion of cells and compared the identified proteins both qualitatively and quantitatively.</p> </sec> <sec id="rcm7230-sec-0002" sec-type="section"> <title>Methods</title> <p>Three hundred cells were directly digested by trypsin, chymotrypsin, or the combination of trypsin and chymotrypsin. The peptides were identified using a LTQ‐Orbitrap XL, and data were analyzed using MaxQuant software.</p> </sec> <sec id="rcm7230-sec-0003" sec-type="section"> <title>Results</title> <p>Different proteases produced different identified protein numbers. Trypsin proved to be the best choice for generating the largest protein number, while other proteases complemented the identification results of trypsin by increasing protein sequence coverage. Concerning the quantitative perspective, using trypsin would produce the biggest number of proteins quantifiable by intensity‐based absolute quantification (iBAQ).</p> </sec> <sec id="rcm7230-sec-0004" sec-type="section"> <title>Conclusions</title> <p>When hundred‐scale cell samples are analyzed, an optimum choice of proteases should be made to realize different analytical objectives. Copyright © 2015 John Wiley & Sons, Ltd.</p> </sec> </abstract>