Detection of functional ricin by immunoaffinity and liquid chromatography-tandem mass spectrometry.
The toxin ricin is a biological weapon that may be used for bioterrorist purposes. As a member of the group of ribosome-inactivating proteins (RIPs), ricin has an A-chain possessing N-glycosidase activity which irreversibly inhibits protein synthesis. In this paper, we demonstrate that provided appropriate sample preparation is used, this enzymatic activity can be exploited for functional ricin detection with sensitivity similar to the best ELISA and specificity allowing application to environmental samples. Ricin is first captured by a monoclonal antibody directed against the B chain and immobilized on magnetic beads. Detection is then realized by determination of the adenine released by the A chain from an RNA template using liquid chromatography coupled to tandem mass spectrometry. The immunoaffinity step combined with the enzymatic activity detection leads to a specific assay for the entire functional ricin with a lower limit of detection of 0.1 ng/mL (1.56 pM) after concentration of the toxin from a 500 microL sample size. The variability of the assay was 10%. Finally, the method was applied successfully to milk and tap or bottled water samples.
Becher
F
F
CEA, Service de Pharmacologie et d'Immunologie, 91191 Gif-sur-Yvette, France and LCSOB UMR 7613 CNRS, Université Pierre et Marie Curie, Paris, France. francois.becher@cea.fr
Duriez
E
E
Volland
H
H
Tabet
J C
JC
Ezan
E
E
D000225:Adenine / Q000032:analysis*; D023482:Bioterrorism; D002846:Chromatography, Affinity / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000737:chemistry*; D012313:RNA / Q000737:chemistry; D015203:Reproducibility of Results; D012276:Ricin / Q000737:chemistry* / Q000276:immunology; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
D012313:RNA; D012276:Ricin; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D000225:Adenine