Assessment of the Reproducibility of Random Hexapeptide Peptide Library-Based Protein Normalization

Dwivedi, Ravi C.; Krokhin, Oleg V.; Cortens, John P.; Wilkins, John A.

doi:10.1021/pr900608z

Cited by 40 publications

(24 citation statements)

References 30 publications

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“…Moreover one-third of the protein spots identified from the CPLL eluate contained fibrinogen. In a way, these results were quite similar to those of Dwivedi et al [14] or to Ernoult et al [15]. This was quite puzzling, since in our hands, and in all of our earlier reports, typically 300-500% more proteins could be identified in PM-treated samples.…”

supporting

confidence: 90%

“…It turns out that there are essentially four main elution cocktails, which are listed here: (i) 4 M urea1 1% CHAPS [13]; (ii) 4 M urea11% CHAPS15% acetic acid [14,15,17,19]; (iii) 8 M urea12% CHAPS15% acetic acid [18,20,23]; and (iv) boiling 4% SDS125 mM DTT [21][22][23]. Briefly, the experimental protocol is as follows: 1 mL of serum is incubated for 3 h, on a rotating platform, with 50 mL of CPLL beads.…”

mentioning

confidence: 99%

“…The results are as follows: eluant 4 M urea11% CHAPS seems to elute not more than 20% of the captured species, the remaining 80% appearing only in the second eluate. In the case of 4 M urea, 1% CHAPS and 5% acetic acid, the elution power was augmented by a factor of three, up to 60%, whereas in the last case (8 M urea, 1% CHAPS and 5% acetic acid) the eluate represents about 80% (4 Â of eluant (i)) of the total [15] (i) 4 M urea, 1% CHAPS, 5% acetic acid; 320 proteins in PM; 332 immuno-depletion (ii) 6 M guanidine HCl, pH 6.0 [17] 4 M urea, 1% CHAPS, 5% acetic acid Many more SELDI peaks than control [18] (i) 8 M urea, 2% CHAPS, 5% acetic acid Identical spots in 2D maps (ii) 7 M urea, 2 M thiourea, 4% CHAPS, 40 mM Tris [19] 4 M urea, 1% CHAPS, 5% acetic acid Control: 157 spots; PM treated: 557 spots, IPGs pH 4-7 [20] 8 M urea, 2% CHAPS, 5% acetic acid Control: 48 peaks; PM-treated: 136 Peaks in SELDI [13] 4 M urea, 1% CHAPS Only high abundance proteins 1 mg to 40 mg/mL [21] 100 mM Tris-HCL pH 6.8, 4% SDS, 10% 2-ME, boil Untreated: 251 proteins; PM-treated: 693 proteins; saliva [14] 33.3% 2-propanol, 16.7% ACN, 0.1% TFA) 108 proteins unique to PM; 100 proteins unique to IgY; 404 total proteins; sera [23] (i) 8 M urea, 2% CHAPS, 5% acetic acid; Control: 211 proteins ; CPLL-treated: 512 a)…”

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confidence: 99%

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“Proteomineering” serum biomarkers. A Study in Scarlet

et al. 2011

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The performance of sera pre-treatment for biomarker searching via combinatorial peptide ligand libraries (CPLL) has recently been challenged (Proteomics 2010, 10, 1416-1425) and stated to allow discovery of only medium to high-abundance proteins. We have thus investigated four elution protocols, as published in recent reports: (i) in 4 M urea+1% CHAPS; (ii) in 4 M urea+1% CHAPS+5% acetic acid; (iii) in 8 M urea+2% CHAPS+5% acetic acid; (iv) in boiling 4% SDS+25 mM DTT. One milliliter of serum, in all cases, was captured with 50 μL of CPLL beads, which were then eluted with the four eluants described above. In the first three cases, after the first elution, the beads were re-eluted with cocktail (iv), known to offer maximal release of proteins adsorbed by the CPLL ligands. Eluant (i) released only ca. 20% of the species adsorbed, eluant (ii) ca. 60%, eluant (iii) ca. 80%. Thus, the poor performance of the CPLL methodology, as reported in (i) is not due to any fault of the capture technique, but simply to the adoption of a very poor elution protocol. Even those using eluants (ii) and (iii) should know that a substantial fraction of the captured species still remains bound to the beads and is thus not available to biomarker discovery. Once more, eluant (iv) is recognized as the only one able to offer optimal recovery from the CPLL baits.

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supporting

confidence: 90%

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confidence: 99%

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confidence: 99%

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“Proteomineering” serum biomarkers. A Study in Scarlet

et al. 2011

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“…However, no differential regulation was observed for these medium and low abundant proteins in our analysis. We also observed high standard deviations for some of the proteins shown in Table 1, which could be explained by the multiple mechanisms attributed to the ProteoMiner bead methodology including (1) selective binding of proteins to highly diverse but specific peptide binding sites, (2) a general hydrophobic interaction (Keidel et al, 2010) proposed for the action of hexapeptide ligand library beads that has been described to contribute and increase variations up to twofold (Dwivedi et al, 2010) in the relative efficiencies of capture of some proteins by equalizer beads in ProteoMiner kit. In this study, we pooled the samples from each cohort prior to the triplicate analysis.…”

Section: Discussionmentioning

confidence: 92%

“…This technology has been applied successfully for the investigation of low abundant plasma and serum proteome (Ye et al, 2010). ProteoMiner equalization was also proved to be a complementary method for quantitative proteomics analysis (Dwivedi et al, 2010) and it was successfully applied to plasma biomarker discovery (Ernoult et al, 2010) and clinical proteomics (Hartwig et al, 2009). By combining the proteominer enrichment and quantitative proteomics, we identified a number of medium to low abundant proteins in addition to several classical plasma proteins, in the analysis of plasma samples from uninfected and HIV-1/HCV-infected patients.…”

Section: Discussionmentioning

confidence: 99%

Investigation of Plasma Biomarkers in HIV-1/HCV Mono- and Coinfected Individuals by Multiplex iTRAQ Quantitative Proteomics

Shetty

Jain

Nickens

et al. 2011

OMICS: A Journal of Integrative Biology

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The analysis of plasma samples from HIV-1/HCV mono-and coinfected individuals by quantitative proteomics is an efficient strategy to investigate changes in protein abundances and to characterize the proteins that are the effectors of cellular functions involved in viral pathogenesis. In this study, the infected and healthy plasma samples (in triplicate) were treated with ProteoMiner beads to equalize protein concentrations and subjected to 4-plex iTRAQ labeling and liquid chromatography/mass spectrometry (LC-MS/MS) analysis. A total of 70 proteins were identified with high confidence in the triplicate analysis of plasma proteins and 65% of the proteins were found to be common among the three replicates. Apolipoproteins and complement proteins are the two major classes of proteins that exhibited differential regulation. The results of quantitative analysis revealed that APOA2, APOC2, APOE, C3, HRG proteins were upregulated in the plasma of all the three HIV-1 mono-, HCV mono-, and coinfected patient samples compared to healthy control samples. Ingenuity pathway analysis (IPA) of the upregulated proteins revealed that they are implicated in the hepatic lipid metabolism, inflammation, and acute-phase response signaling pathways. Thus, we identified several differentially regulated proteins in HIV-1/HCV mono and coinfected plasma samples that may be potential biomarkers for liver disease.

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Combinatorial peptide ligand library plasma treatment: Advantages for accessing low‐abundance proteins

et al. 2010

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Depletion of major blood proteins is one of the most promising approaches to accessing low-abundance biomarkers. This study compared the use of combinatorial peptide ligand library (CPLL) and albumin and immunoglobulins (IgGs) depletion technology for accessing these low-abundance proteins in plasma using 2-DE in an acidic restricted pH range (4-7). Compared with native plasma, both techniques enlarge the visibility of other proteins than albumin and IgG, but there were marked differences in their composition. An increase of the number of spots was detected compared with native plasma (157 spots) with 427 and 557 spots, respectively, detected with albumin and IgG depletion, and CPLL treatment. We selected 70 spots to be identified by MALDI-TOF related to their absence in the 2-D gels from native or albumin and IgG-depleted plasma. The 42 spots identified corresponded to 24 different proteins, with more than half of the proteins which did not belong to the major plasma proteins. CPLL treatment allowed the accessibility to proteolytic fragments obtained from major plasma proteins. We found a large superiority of the CPLL approach over the albumin and IgG depletion process. These findings show the utility of depleting major blood proteins to be able to access low-abundance proteins and the potential of CPLL to select and identify candidate biomarkers in clinical studies.

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Assessment of the Reproducibility of Random Hexapeptide Peptide Library-Based Protein Normalization

Cited by 40 publications

References 30 publications

“Proteomineering” serum biomarkers. A Study in Scarlet

“Proteomineering” serum biomarkers. A Study in Scarlet

Investigation of Plasma Biomarkers in HIV-1/HCV Mono- and Coinfected Individuals by Multiplex iTRAQ Quantitative Proteomics

Combinatorial peptide ligand library plasma treatment: Advantages for accessing low‐abundance proteins

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