Universität zu Köln

Reichel, Christian, Benetka, Wolfgang, Lorenc, Barbara and Thevis, Mario (2016). Evaluation of AMGEN clone 9G8A anti-Epo antibody for application in doping control. Drug Test. Anal., 8 (11-12). S. 1131 - 1138. HOBOKEN: WILEY-BLACKWELL. ISSN 1942-7611

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Abstract

The two mouse monoclonal anti-erythropoietin (EPO) antibodies clone AE7A5 (generated by using a 26 amino acid N-terminal EPO-peptide) and 9G8A (developed by immunizing mice with full length human EPO) are both directed against linear epitopes at the N-terminus of EPO. While AE7A5 has been commercially available for many years, 9G8A was made for Amgen's internal research purposes. In the past, the commercial antibody was shown to cross-react with several proteins unrelated to EPO (e.g. E. coli thioredoxin reductase, zinc alpha 2glycoprotein, S. cerevisiae enolase, human neuron-specific enolase, and human non-neuronal enolase). However, it displayed high sensitivity for detecting recombinant EPO (rEPO) misuse by athletes on Western blots. We evaluated the potential use of clone 9G8A for doping control purposes. While 9G8A showed lower sensitivity than AE7A5 (ca 45% on isoelectric focusing (IEF)-polyacrylamide gel electrophoresis (PAGE), ca 40% on sodium dodecyl sulfate (SDS)-and sarcosyl (SAR)PAGE), non-specific binding of the five proteins was not observed. The cross-reactivity of AE7A5 can be overcome by immunoaffinity purification of EPO before electrophoresis and Western blotting. Similar to AE7A5, clone 9G8A is also suited for Western double-blotting. Copyright (C) 2016 John Wiley & Sons, Ltd.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Reichel, Christian UNSPECIFIED UNSPECIFIED UNSPECIFIED Benetka, Wolfgang UNSPECIFIED UNSPECIFIED UNSPECIFIED Lorenc, Barbara UNSPECIFIED UNSPECIFIED UNSPECIFIED Thevis, Mario UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-256172
DOI:	10.1002/dta.2057
Journal or Publication Title:	Drug Test. Anal.
Volume:	8
Number:	11-12
Page Range:	S. 1131 - 1138
Date:	2016
Publisher:	WILEY-BLACKWELL
Place of Publication:	HOBOKEN
ISSN:	1942-7611
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language RECOMBINANT-HUMAN-ERYTHROPOIETIN; 2-DIMENSIONAL GEL-ELECTROPHORESIS; URINE; PAGE; BINDING; PERFORMANCE; AE7A5 Multiple languages Biochemical Research Methods; Chemistry, Analytical; Pharmacology & Pharmacy Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/25617