Preanalytical and analytical variation of surface-enhanced laser desorption-ionization time-of-flight mass spectrometry of human serum

Albrethsen, Jakob; Bøgebo, Rikke; Olsen, John Elmerdahl; Raskov, Hans; Gammeltoft, Steen

doi:10.1515/cclm.2006.228

Cited by 57 publications

(50 citation statements)

References 31 publications

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“…Protein m/z 2743, 3891, 4282, 10838 and 16120 were shown to be useful in classifying RCC and HC in classification trees with a sensitivity and specificity ranging from 62 to 85% (both independent validation and crossvalidation). The intra-assay CV of our peak intensities, including the discriminating peaks, was higher than what is generally reported (~20-30%) (36,37). This might be caused by the manual instead of automated processing of the samples.…”

Section: Discussioncontrasting

confidence: 70%

Identification of two new serum protein profiles for renal cell carcinoma

Engwegen

Mehra²,

Haanen³

et al. 2009

Oncol Rep

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Abstract. Renal cell carcinoma (RCC) is the most lethal urological cancer, and survival greatly depends on early diagnosis. Therefore, reliable, new biomarkers for detection of RCC are required. We assessed serum protein profiles of samples from two institutes with SELDI-TOF MS in duplicate on CM10 chips at pH 6.0 (set 1: 37 RCC + 32 healthy controls (HC), set 2: 20 RCC + 25 HC). Mean peak intensities of detected proteins were compared between RCC and HC with non-parametric testing. Classification trees were built with discriminating peaks using one sample set as training set and the other as independent validation set. We found 15 peaks significantly different (p<0.01) between RCC and HC. Two classification trees could be built with these peaks. Tree A achieved 75% sensitivity and 85% specificity for crossvalidation and 76 and 65% for independent validation. Tree B had 71 and 62% sensitivity and specificity for cross-validation and 83 and 82% for independent validation. Although two serum protein profiles comprising 5 protein peaks were found that could separate RCC from HC, the sensitivity and specificity is not sufficient to recommend large scale use. Upon structural identification and quantitative validation, however, these proteins might prove suitable markers in the follow up of RCC patients.

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Section: Discussioncontrasting

confidence: 70%

Identification of two new serum protein profiles for renal cell carcinoma

Engwegen

Mehra²,

Haanen³

et al. 2009

Oncol Rep

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show abstract

“…The SELDI-TOF MS instrument is Clinical Chemistry 53, No. 5,2007 designed to produce a reproducible protein profile over a relatively wide mass range at the price of lower mass accuracy and lower resolution of the spectral output. With the PBS II SELDI-TOF MS instrument, the mean interlaboratory CV of the m/z value is Ͻ0.1% (16 ), whereas the latest SELDI model (the Protein Reader Series 4000) has improved resolution (27 ).…”

Section: Approaches To Improve the Analytical Performance Of Maldi Prmentioning

confidence: 99%

Reproducibility in Protein Profiling by MALDI-TOF Mass Spectrometry

2007

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Background: Protein profiling with high-throughput sample preparation and MALDI-TOF MS analysis is a new potential tool for diagnosis of human diseases. However, analytical reproducibility is a significant challenge in MALDI protein profiling. This minireview summarizes studies of reproducibility of MALDI protein profiling and current approaches to improve its analytical performance. Methods: The PubMed database was searched using combinations of the following search terms: MALDI, SELDI, reproducibility, variation, precision, peak intensity, quantification, peptide, biomarkers, and proteomics. Acceptance criteria were detailed reports on the reproducibility with MALDI protein profiling and studies describing efforts to improve the analytical performance with this technology. Results: The reported intraexperiment CVs of the peak intensity vary highly between individual protein peaks, with the reported mean CV of the peak intensity varying among studies from 4% to 26%. There is additional interexperiment variation in peak intensity. Current approaches to improve the analytical performance of MALDI protein profiling include automated sample processing, extensive prefractionation strategies, immunocapture, prestructured target surfaces, standardized matrix (co)crystallization, improved MALDI-TOF MS instrument components, internal standard peptides, quality-control samples, replicate measurements, and algorithms for normalization and peak detection. Conclusions: Further evaluation and optimization of MALDI-TOF MS is recommended before use in routine analysis.

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“…on analytical outcome, particularly for protein and proteomic studies (3,(8)(9)(10)(11). A recent perspective on the subject has been put forward with specific focus on proteome analyses of blood (12).…”

Section: Introductionmentioning

confidence: 99%

Thrombin induces broad spectrum proteolysis in human serum samples

O'Mullan¹,

Craft²,

Yi³

et al. 2009

Clinical Chemistry and Laboratory Medicine

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Background: During clotting, a thrombin cleaves fibrinogen releasing fibrinopeptide A (FPA). FPA is easily identified in serum using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). Using MALDI-TOF MS, we observed multiple, progressively shorter fragments of serum FPA. Following ambient incubation of serum, variations in the content of FPA fragments occur over time. Denaturation of a thrombin by heating the serum sample appears to minimize this variation. These observations suggest that intrinsic proteolytic and peptidolytic activity is elevated in serum and perhaps originates from the coagulation cascade enzymes themselves, especially a thrombin. Methods: Extrinsic addition of a thrombin to a subset (3-30 kDa) of plasma proteins was carried out to induce proteolysis and to examine the resultant peptides to reveal a thrombin susceptible parent proteins. One of these identified proteins, hemopexin, was directly digested by a thrombin and the peptides examined to confirm the observations from the initial plasma protein digestion. Results: Extrinsic addition of a thrombin to a subset (3-30 kDa) of plasma proteins results in wide-spread digestion of proteins unrelated to coagulation, revealing a substrate range encompassing more than fibrinogen. Direct digestion of one of these proteins, hemopexin, by a thrombin confirms these observations. Conclusions: The resulting peptides indicate broad tolerance beyond the consensus R-G cleavage site of fibrinogen; in fact, there appears to be no bias for the amino acid following the R/K residue. These data support our hypothesis that the enzymatic activities inherent to coagulation, or at least to thrombin, contribute to destabilization of the protein and peptide content of serum. Clin Chem Lab Med 2009;47:685-93.

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Preanalytical and analytical variation of surface-enhanced laser desorption-ionization time-of-flight mass spectrometry of human serum

Cited by 57 publications

References 31 publications

Identification of two new serum protein profiles for renal cell carcinoma

Identification of two new serum protein profiles for renal cell carcinoma

Reproducibility in Protein Profiling by MALDI-TOF Mass Spectrometry

Thrombin induces broad spectrum proteolysis in human serum samples

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