Publication

Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum

Wilffert, D., Reis, C. R., Hermans, J., Govorukhina, N., Tomar, T., de Jong, S., Quax, W. J., van de Merbel, N. C. & Bischoff, R., 19-Nov-2013, In : Analytical Chemistry. 85, 22, p. 10754-10760 7 p.

Research output: Contribution to journalArticleAcademicpeer-review

APA

Wilffert, D., Reis, C. R., Hermans, J., Govorukhina, N., Tomar, T., de Jong, S., Quax, W. J., van de Merbel, N. C., & Bischoff, R. (2013). Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum. Analytical Chemistry, 85(22), 10754-10760. https://doi.org/10.1021/ac4017902

Author

Wilffert, Daniel ; Reis, C.R. ; Hermans, Jos ; Govorukhina, Natalia ; Tomar, Tushar ; de Jong, Steven ; Quax, Wim J. ; van de Merbel, Nico C. ; Bischoff, Rainer. / Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum. In: Analytical Chemistry. 2013 ; Vol. 85, No. 22. pp. 10754-10760.

Harvard

Wilffert, D, Reis, CR, Hermans, J, Govorukhina, N, Tomar, T, de Jong, S, Quax, WJ, van de Merbel, NC & Bischoff, R 2013, 'Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum', Analytical Chemistry, vol. 85, no. 22, pp. 10754-10760. https://doi.org/10.1021/ac4017902

Standard

Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum. / Wilffert, Daniel; Reis, C.R.; Hermans, Jos; Govorukhina, Natalia; Tomar, Tushar; de Jong, Steven; Quax, Wim J.; van de Merbel, Nico C.; Bischoff, Rainer.

In: Analytical Chemistry, Vol. 85, No. 22, 19.11.2013, p. 10754-10760.

Research output: Contribution to journalArticleAcademicpeer-review

Vancouver

Wilffert D, Reis CR, Hermans J, Govorukhina N, Tomar T, de Jong S et al. Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum. Analytical Chemistry. 2013 Nov 19;85(22):10754-10760. https://doi.org/10.1021/ac4017902


BibTeX

@article{03339620fdb64b3bad0863a3cd6b5194,
title = "Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum",
abstract = "The major challenge in targeted protein quantification by LC-MS/MS in serum lies in the complexity of the biological matrix with regard to the wide diversity of proteins and their extremely large dynamic concentration range. In this study, an LC-MS/MS method was developed for the simultaneous quantification of the 60-kDa biopharmaceutical proteins recombinant human tumor necrosis factor-related apoptosis-inducing ligand wild type (rhTRAIL(WT)) and its death receptor 4 (DR4)-specific variant rhTRAIL(4C7) in human and mouse serum. Selective enrichment of TRAIL was accomplished by immobilized metal affinity chromatography (IMAC), which was followed by tryptic digestion of the enriched sample and quantification of a suitable signature peptide. For absolute quantification, (15)N-metabolically labeled internal standards of rhTRAIL(WT) and rhTRAIL(4C7) were used. Since the signature peptides that provided the highest sensitivity and allowed discrimination between rhTRAIL(WT) and rhTRAIL(4C7) contained methionine residues, we oxidized these quantitatively to their sulfoxides by the addition of 0.25% (w/w) hydrogen peroxide. The final method has a lower limit of quantification of 20 ng/mL (ca. 350 pM) and was fully validated according to current international guidelines for bioanalysis. To show the applicability of the LC-MS/MS method for pharmacokinetic studies, we quantified rhTRAIL(WT) and rhTRAIL(4C7) simultaneously in serum from mice injected intraperitoneally at a dose of 5 mg/kg for each protein. This is the first time that two variants of rhTRAIL differing by only a few amino acids have been analyzed simultaneously in serum, an approach that is not possible by conventional enzyme-linked immuno-sorbent assay (ELISA) analysis.",
keywords = "APOPTOSIS-INDUCING LIGAND, MASS-SPECTROMETRY, CANCER, CHROMATOGRAPHY, PROTEOMICS, PROTEINS, TRAIL, DEATH, RECEPTORS, STANDARDS",
author = "Daniel Wilffert and C.R. Reis and Jos Hermans and Natalia Govorukhina and Tushar Tomar and {de Jong}, Steven and Quax, {Wim J.} and {van de Merbel}, {Nico C.} and Rainer Bischoff",
year = "2013",
month = nov,
day = "19",
doi = "10.1021/ac4017902",
language = "English",
volume = "85",
pages = "10754--10760",
journal = "Analytical Chemistry",
issn = "0003-2700",
publisher = "AMER CHEMICAL SOC INC",
number = "22",

}

RIS

TY - JOUR

T1 - Antibody-free LC-MS/MS quantification of rhTRAIL in human and mouse serum

AU - Wilffert, Daniel

AU - Reis, C.R.

AU - Hermans, Jos

AU - Govorukhina, Natalia

AU - Tomar, Tushar

AU - de Jong, Steven

AU - Quax, Wim J.

AU - van de Merbel, Nico C.

AU - Bischoff, Rainer

PY - 2013/11/19

Y1 - 2013/11/19

N2 - The major challenge in targeted protein quantification by LC-MS/MS in serum lies in the complexity of the biological matrix with regard to the wide diversity of proteins and their extremely large dynamic concentration range. In this study, an LC-MS/MS method was developed for the simultaneous quantification of the 60-kDa biopharmaceutical proteins recombinant human tumor necrosis factor-related apoptosis-inducing ligand wild type (rhTRAIL(WT)) and its death receptor 4 (DR4)-specific variant rhTRAIL(4C7) in human and mouse serum. Selective enrichment of TRAIL was accomplished by immobilized metal affinity chromatography (IMAC), which was followed by tryptic digestion of the enriched sample and quantification of a suitable signature peptide. For absolute quantification, (15)N-metabolically labeled internal standards of rhTRAIL(WT) and rhTRAIL(4C7) were used. Since the signature peptides that provided the highest sensitivity and allowed discrimination between rhTRAIL(WT) and rhTRAIL(4C7) contained methionine residues, we oxidized these quantitatively to their sulfoxides by the addition of 0.25% (w/w) hydrogen peroxide. The final method has a lower limit of quantification of 20 ng/mL (ca. 350 pM) and was fully validated according to current international guidelines for bioanalysis. To show the applicability of the LC-MS/MS method for pharmacokinetic studies, we quantified rhTRAIL(WT) and rhTRAIL(4C7) simultaneously in serum from mice injected intraperitoneally at a dose of 5 mg/kg for each protein. This is the first time that two variants of rhTRAIL differing by only a few amino acids have been analyzed simultaneously in serum, an approach that is not possible by conventional enzyme-linked immuno-sorbent assay (ELISA) analysis.

AB - The major challenge in targeted protein quantification by LC-MS/MS in serum lies in the complexity of the biological matrix with regard to the wide diversity of proteins and their extremely large dynamic concentration range. In this study, an LC-MS/MS method was developed for the simultaneous quantification of the 60-kDa biopharmaceutical proteins recombinant human tumor necrosis factor-related apoptosis-inducing ligand wild type (rhTRAIL(WT)) and its death receptor 4 (DR4)-specific variant rhTRAIL(4C7) in human and mouse serum. Selective enrichment of TRAIL was accomplished by immobilized metal affinity chromatography (IMAC), which was followed by tryptic digestion of the enriched sample and quantification of a suitable signature peptide. For absolute quantification, (15)N-metabolically labeled internal standards of rhTRAIL(WT) and rhTRAIL(4C7) were used. Since the signature peptides that provided the highest sensitivity and allowed discrimination between rhTRAIL(WT) and rhTRAIL(4C7) contained methionine residues, we oxidized these quantitatively to their sulfoxides by the addition of 0.25% (w/w) hydrogen peroxide. The final method has a lower limit of quantification of 20 ng/mL (ca. 350 pM) and was fully validated according to current international guidelines for bioanalysis. To show the applicability of the LC-MS/MS method for pharmacokinetic studies, we quantified rhTRAIL(WT) and rhTRAIL(4C7) simultaneously in serum from mice injected intraperitoneally at a dose of 5 mg/kg for each protein. This is the first time that two variants of rhTRAIL differing by only a few amino acids have been analyzed simultaneously in serum, an approach that is not possible by conventional enzyme-linked immuno-sorbent assay (ELISA) analysis.

KW - APOPTOSIS-INDUCING LIGAND

KW - MASS-SPECTROMETRY

KW - CANCER

KW - CHROMATOGRAPHY

KW - PROTEOMICS

KW - PROTEINS

KW - TRAIL

KW - DEATH

KW - RECEPTORS

KW - STANDARDS

U2 - 10.1021/ac4017902

DO - 10.1021/ac4017902

M3 - Article

C2 - 24125577

VL - 85

SP - 10754

EP - 10760

JO - Analytical Chemistry

JF - Analytical Chemistry

SN - 0003-2700

IS - 22

ER -

ID: 5995285