Quantification of Target Peptides or Proteins by Liquid Chromatography Mass Spectrometry with Multiple Reaction Monitoring

JIA Xiao-Fang, LIU Yong-Fu, TENG Zhen-Lin, YIN Lin, LIU Bao-Chi, ZHANG Li-Jun

Chinese Journal of Biochemistry and Molecular Biology ›› 2012, Vol. 28 ›› Issue (1) : 86-92.

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Chinese Journal of Biochemistry and Molecular Biology ›› 2012, Vol. 28 ›› Issue (1) : 86-92.
Techniques and Methods

Quantification of Target Peptides or Proteins by Liquid Chromatography Mass Spectrometry with Multiple Reaction Monitoring

  • LIU Yong-Fu1),2),JIA Xiao-Fang1),TENG Zhen-Lin1),YIN Lin1),LIU Bao-Chi1), ZHANG Li-Jun1),3) *
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Abstract

This study was to establish a modified plasma peptide extraction method and to develop a mass spectrometry quantification method for target peptides or proteins. Three techniques of ultrafiltration, organic solvent extraction and solid phase extraction were used to extract peptides from plasma samples. Tricine-SDS-PAGE was used to determine the extraction efficiency. Peptide and protein quantification were completed by liquid chromatography combined with mass spectrometry in multiple reaction monitoring (MRM) mode. The Tricine-SDS-PAGE results showed that acetonitrile (ACN) precipitation was the most efficient approach for low molecular plasma peptides enrichment besides for excluding the high molecular protein contaminations. The correlation coefficients of the standard curves were 0.995 and 0.999 in the quantification of peptide or protein standards by liquid chromatographymass spectrometry (LC-MS), respectively. In conclusion, we successfully established a simple and efficient plasma peptide extraction method and a subsequent LC-MS-MRM method for peptide and protein quantifications. These methods might be applied to the quantification of target peptides or proteins in complex samples.

Key words

peptide / protein / multiple reaction monitoring (MRM) / quantification / liquid chromatography mass spectrometry(LC-MS)

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JIA Xiao-Fang, LIU Yong-Fu, TENG Zhen-Lin, YIN Lin, LIU Bao-Chi, ZHANG Li-Jun. Quantification of Target Peptides or Proteins by Liquid Chromatography Mass Spectrometry with Multiple Reaction Monitoring[J]. Chinese Journal of Biochemistry and Molecular Biology, 2012, 28(1): 86-92

References

[1] Petricoin E F, Belluco C, Araujo R P, et al. The blood peptidome: a higher dimension of information content for cancer biomarker discovery[J]. Nat Rev Cancer, 2006, 6 (12):961-967
[2] Liu Y, Uboh C E, Soma L R, et al. Efficient use of retention time for the analysis of 302 drugs in equine plasma by liquid chromatography-MS/MS with scheduled multiple reaction monitoring and instant library searching for doping control[J]. Anal Chem, 2011, 83 (17):6834-6841
[3] Lee J H, Lim H, Kim D G, et al. Quantification of oltipraz using liquid chromatography-tandem mass spectrometry and its application to a pharmacokinetic study in rat plasma[J]. J Pharm Biomed Anal, 2011, 56 (3):623-626
[4] Ang C S, Nice E C. Targeted in-gel MRM: a hypothesis driven approach for colorectal cancer biomarker discovery in human feces[J]. J Proteome Res, 2010, 9 (9):4346-4355
[5] Kuzyk M A, Smith D, Yang J, et al. Multiple reaction monitoring-based, multiplexed, absolute quantitation of 45 proteins in human plasma[J]. Mol Cell Proteomics, 2009, 8 (8):1860-1877
[6] 赵焱, 应万涛, 钱小红. 质谱MRM技术在蛋白质组学研究中的应用[J]. 生命的化学(Zhao Yan, Ying Wan-Tao, Qian Xiao-Hong. Application of mass spectrometry MRM technology in proteome research[J].Chem Life), 2008, 28 (2):210-213
[7] Ueda K, Saichi N, Takami S, et al. A comprehensive peptidome profiling technology for the identification of early detection biomarkers for lung adenocarcinoma[J]. PLoS One, 2011, 6 (4): e18567
[8] Sch?gger H. Tricine-SDS-PAGE[J]. Nat Protoc, 2006, 1 (1):16-22
[9] Zhang L, Jia X, Zhang X, et al. Alpha-1 antitrypsin variants in plasma from HIV-infected patients revealed by proteomic and glycoproteomic analysis[J]. Electrophoresis, 2010, 31 (20): 3437-3445
[10] Schulz-Knappe P, Zucht H D, Heine G, et al. Peptidomics: the comprehensive analysis of peptides in complex biological mixtures[J]. Comb Chem High Throughput Screen, 2001, 4 (2): 207-217
[11] Jiang X, Ye M, Zou H. Technologies and methods for sample pretreatment in efficient proteome and peptidome analysis[J]. Proteomics, 2008, 8 (4):686-705
[12] Aristoteli L P, Molloy M P, Baker M S. Evaluation of endogenous plasma peptide extraction methods for mass spectrometric biomarker discovery[J]. J Proteome Res, 2007, 6 (2):571-581
[13] Zheng X, Baker H, Hancock W. Analysis of the low molecular weight serum peptidome using ultrafiltration and a hybrid ion trap-Fourier transform mass spectrometer[J]. J Chromatogr A, 2006, 1120 (1-2):173-184
[14] Zougman A, Pilch B, Podtelejnikov A, et al. Integrated analysis of the cerebrospinal fluid peptidome and proteome[J]. J Proteome Res, 2008, 7 (1):386-399
[15] Fiedler G M, Baumann S, Leichtle A, et al. Standardized peptidome profiling of human urine by magnetic bead separation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry[J]. Clin Chem, 2007, 53 (3):421-428
[16] Villanueva J, Philip J, Entenberg D, et al. Serum peptide profiling by magnetic particle-assisted, automated sample processing and MALDI-TOF mass spectrometry[J]. Anal Chem, 2004, 76 (6): 1560-1570
[17] Tucholska M, Scozzaro S, Williams D, et al. Endogenous peptides from biophysical and biochemical fractionation of serum analyzed by matrix-assisted laser desorption/ionization and electrospray ionization hybrid quadrupole time-of-flight[J].Anal Biochem,2007,370(2):228- 245
[18] Chertov O, Biragyn A, Kwak L W, et al. Organic solvent extraction of proteins and peptides from serum as an effective sample preparation for detection and identification of biomarkers by mass spectrometry[J]. Proteomics, 2004, 4 (4):1195-1203
[19] Bertile F, Robert F, Delval-Dubois V, et al. Endogenous plasma peptide detection and identification in the rat by a combination of fractionation methods and mass spectrometry[J]. Biomark Insights, 2007, 2:385-401


Funding

Supported by Postdoctoral Science Foundation of China (No.20100471000,No.201104484),Wang BaoEn Liver Fibrosis Research Foundation from Chinese Foundation for Hepatitis Prevention and Control (No.20100031)

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