SARCOSYL-PAGE: Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins

Publikationen: Beitrag in Buch/Bericht/KonferenzbandBeiträge in SammelwerkenForschungBegutachtung

Standard

SARCOSYL-PAGE : Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins. / Reichel, Christian; Gmeiner, Günter; Reihlen, Philipp; Thevis, Mario; Schänzer, Wilhelm.

Electrophoretic separation of proteins: methods and protocols. Hrsg. / Biji T. Kurien; R. Hal Scofield. New York : Humana Press, 2019. S. 131-149 (Methods in Molecular Biology; Band 1855).

Publikationen: Beitrag in Buch/Bericht/KonferenzbandBeiträge in SammelwerkenForschungBegutachtung

Harvard

Reichel, C, Gmeiner, G, Reihlen, P, Thevis, M & Schänzer, W 2019, SARCOSYL-PAGE: Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins. in BT Kurien & RH Scofield (Hrsg.), Electrophoretic separation of proteins: methods and protocols. Methods in Molecular Biology, Bd. 1855, Humana Press, New York, S. 131-149. https://doi.org/10.1007/978-1-4939-8793-1_14

APA

Reichel, C., Gmeiner, G., Reihlen, P., Thevis, M., & Schänzer, W. (2019). SARCOSYL-PAGE: Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins. in B. T. Kurien, & R. H. Scofield (Hrsg.), Electrophoretic separation of proteins: methods and protocols (S. 131-149). (Methods in Molecular Biology; Band 1855). Humana Press. https://doi.org/10.1007/978-1-4939-8793-1_14

Vancouver

Reichel C, Gmeiner G, Reihlen P, Thevis M, Schänzer W. SARCOSYL-PAGE: Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins. in Kurien BT, Scofield RH, Hrsg., Electrophoretic separation of proteins: methods and protocols. New York: Humana Press. 2019. S. 131-149. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-4939-8793-1_14

Bibtex

@inbook{ae75c6f7b8c347eead105a60e95e7995,
title = "SARCOSYL-PAGE: Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins",
abstract = "PEGylation of recombinant proteins and synthetic peptides aims to generate biopharmaceuticals with altered physical properties. The modification may lead to a prolonged serum half-life caused by decreased receptor-mediated endocytosis and/or delay in renal clearance caused by the increased hydrodynamic volume of the pharmaceutical. MIRCERA, a PEGylated recombinant erythropoietin (rhEPO) used in the treatment of anemia due to chronic kidney disease, has also been abused by athletes as performance-enhancing drug. While it can be detected by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, the sensitivity of the test is significantly lower compared to other epoetins. By replacing SDS with sarcosyl in the sample and running buffers, the interaction between SDS and the PEG group of the protein no longer reduces the affinity of the monoclonal anti-EPO antibody (clone AE7A5) to the protein chain. Contrary to SDS, sarcosyl only binds to the amino acid chain of the PEGylated protein and thus leads to a sharper electrophoretic band and enhanced antibody binding. While the method was originally developed for anti-doping purposes, it may also be useful for the electrophoretic separation and immunological detection of other PEGylated proteins. Protocols for urine and serum are presented. They are also applicable for the general detection of EPO-based erythropoiesis-stimulating agents (ESA) in these matrices.",
keywords = "BlotCycler, Doping control, Erythropoietin (EPO), Immunoblotting, MIRCERA, PEGylated proteins, SAR-PAGE, SDS-PAGE, Sarcosyl",
author = "Christian Reichel and G{\"u}nter Gmeiner and Philipp Reihlen and Mario Thevis and Wilhelm Sch{\"a}nzer",
year = "2019",
doi = "10.1007/978-1-4939-8793-1_14",
language = "English",
isbn = "978-1-4939-8792-4 ",
series = "Methods in Molecular Biology",
publisher = "Humana Press",
pages = "131--149",
editor = "Kurien, {Biji T.} and Scofield, {R. Hal}",
booktitle = "Electrophoretic separation of proteins",
address = "United States",

}

RIS

TY - CHAP

T1 - SARCOSYL-PAGE

T2 - Optimized Protocols for the Separation and Immunological Detection of PEGylated Proteins

AU - Reichel, Christian

AU - Gmeiner, Günter

AU - Reihlen, Philipp

AU - Thevis, Mario

AU - Schänzer, Wilhelm

PY - 2019

Y1 - 2019

N2 - PEGylation of recombinant proteins and synthetic peptides aims to generate biopharmaceuticals with altered physical properties. The modification may lead to a prolonged serum half-life caused by decreased receptor-mediated endocytosis and/or delay in renal clearance caused by the increased hydrodynamic volume of the pharmaceutical. MIRCERA, a PEGylated recombinant erythropoietin (rhEPO) used in the treatment of anemia due to chronic kidney disease, has also been abused by athletes as performance-enhancing drug. While it can be detected by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, the sensitivity of the test is significantly lower compared to other epoetins. By replacing SDS with sarcosyl in the sample and running buffers, the interaction between SDS and the PEG group of the protein no longer reduces the affinity of the monoclonal anti-EPO antibody (clone AE7A5) to the protein chain. Contrary to SDS, sarcosyl only binds to the amino acid chain of the PEGylated protein and thus leads to a sharper electrophoretic band and enhanced antibody binding. While the method was originally developed for anti-doping purposes, it may also be useful for the electrophoretic separation and immunological detection of other PEGylated proteins. Protocols for urine and serum are presented. They are also applicable for the general detection of EPO-based erythropoiesis-stimulating agents (ESA) in these matrices.

AB - PEGylation of recombinant proteins and synthetic peptides aims to generate biopharmaceuticals with altered physical properties. The modification may lead to a prolonged serum half-life caused by decreased receptor-mediated endocytosis and/or delay in renal clearance caused by the increased hydrodynamic volume of the pharmaceutical. MIRCERA, a PEGylated recombinant erythropoietin (rhEPO) used in the treatment of anemia due to chronic kidney disease, has also been abused by athletes as performance-enhancing drug. While it can be detected by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, the sensitivity of the test is significantly lower compared to other epoetins. By replacing SDS with sarcosyl in the sample and running buffers, the interaction between SDS and the PEG group of the protein no longer reduces the affinity of the monoclonal anti-EPO antibody (clone AE7A5) to the protein chain. Contrary to SDS, sarcosyl only binds to the amino acid chain of the PEGylated protein and thus leads to a sharper electrophoretic band and enhanced antibody binding. While the method was originally developed for anti-doping purposes, it may also be useful for the electrophoretic separation and immunological detection of other PEGylated proteins. Protocols for urine and serum are presented. They are also applicable for the general detection of EPO-based erythropoiesis-stimulating agents (ESA) in these matrices.

KW - BlotCycler

KW - Doping control

KW - Erythropoietin (EPO)

KW - Immunoblotting

KW - MIRCERA

KW - PEGylated proteins

KW - SAR-PAGE

KW - SDS-PAGE

KW - Sarcosyl

UR - https://www.mendeley.com/catalogue/ab2c2389-4c52-3d80-831d-758252984a38/

U2 - 10.1007/978-1-4939-8793-1_14

DO - 10.1007/978-1-4939-8793-1_14

M3 - Contributions to collected editions/anthologies

C2 - 30426415

SN - 978-1-4939-8792-4

T3 - Methods in Molecular Biology

SP - 131

EP - 149

BT - Electrophoretic separation of proteins

A2 - Kurien, Biji T.

A2 - Scofield, R. Hal

PB - Humana Press

CY - New York

ER -

ID: 4726351