title: Qualitative and Quantitative Detection of Botulinum Neurotoxins from Complex Matrices: Results of the First International Proficiency Test.
abstract: In the framework of the EU project EQuATox, a first international proficiency test (PT) on the detection and quantification of botulinum neurotoxins (BoNT) was conducted. Sample materials included BoNT serotypes A, B and E spiked into buffer, milk, meat extract and serum. Different methods were applied by the participants combining different principles of detection, identification and quantification. Based on qualitative assays, 95% of all results reported were correct. Successful strategies for BoNT detection were based on a combination of complementary immunological, MS-based and functional methods or on suitable functional in vivo/in vitro approaches (mouse bioassay, hemidiaphragm assay and Endopep-MS assay). Quantification of BoNT/A, BoNT/B and BoNT/E was performed by 48% of participating laboratories. It turned out that precise quantification of BoNT was difficult, resulting in a substantial scatter of quantitative data. This was especially true for results obtained by the mouse bioassay which is currently considered as "gold standard" for BoNT detection. The results clearly demonstrate the urgent need for certified BoNT reference materials and the development of methods replacing animal testing. In this context, the BoNT PT provided the valuable information that both the Endopep-MS assay and the hemidiaphragm assay delivered quantitative results superior to the mouse bioassay.
authors.affiliation: Biological Toxins, Centre for Biological Threats and Special Pathogens, Robert Koch Institute, Seestrasse 10, 13353 Berlin, Germany. worbss@rki.de.
authors.forename: Sylvia
authors.initials: S
authors.lastname: Worbs
authors.affiliation: Biological Toxins, Centre for Biological Threats and Special Pathogens, Robert Koch Institute, Seestrasse 10, 13353 Berlin, Germany. fiebigu@rki.de.
authors.forename: Uwe
authors.initials: U
authors.lastname: Fiebig
authors.affiliation: European Commission, Joint Research Centre, Institute for Reference Materials and Measurements, Retieseweg 111, 2440 Geel, Belgium. reinhard.zeleny@ec.europa.eu.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Zeleny
authors.affiliation: European Commission, Joint Research Centre, Institute for Reference Materials and Measurements, Retieseweg 111, 2440 Geel, Belgium. heinz.schimmel@ec.europa.eu.
authors.forename: Heinz
authors.initials: H
authors.lastname: Schimmel
authors.affiliation: toxogen GmbH, Feodor-Lynen-Strasse 35, 30625 Hannover, Germany. rummel@toxogen.de.
authors.forename: Andreas
authors.initials: A
authors.lastname: Rummel
authors.affiliation: ChemStat, Aarstrasse 98, 3005 Bern, Switzerland. info@chemstat.ch.
authors.forename: Werner
authors.initials: W
authors.lastname: Luginbühl
authors.affiliation: Biological Toxins, Centre for Biological Threats and Special Pathogens, Robert Koch Institute, Seestrasse 10, 13353 Berlin, Germany. dornerb@rki.de.
authors.forename: Brigitte G
authors.initials: BG
authors.lastname: Dorner
mesh_terms: D000818:Animals; D001905:Botulinum Toxins / Q000032:analysis* / Q000633:toxicity; D019274:Botulinum Toxins, Type A / Q000032:analysis* / Q000633:toxicity; D002021:Buffers; D005506:Food Contamination / Q000032:analysis; D006801:Humans; D059021:Laboratory Proficiency Testing; D007928:Lethal Dose 50; D008460:Meat / Q000032:analysis; D051379:Mice; D008892:Milk / Q000737:chemistry; D009498:Neurotoxins / Q000032:analysis* / Q000633:toxicity; D044967:Serum / Q000737:chemistry; D012710:Serum Albumin, Bovine / Q000737:chemistry
chemical_list: D002021:Buffers; D009498:Neurotoxins; C096323:rimabotulinumtoxinB; D012710:Serum Albumin, Bovine; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; C545476:incobotulinumtoxinA; C096326:botulinum toxin type E
keywords: BoNT; proficiency test; reference material; standardized detection