Detection of Ricin in Complex Samples by Immunocapture and Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

Abstract

Ricin, the toxin component of Ricinus communis is considered as a potential chemical weapon. Several complementary techniques are required to confirm its presence in environmental samples. Here, we report a method combining immunocapture and analysis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the accurate detection of different species of R. communis. Liquid environmental samples were applied to magnetic particles coated with a monoclonal antibody directed against the B-chain of the toxin. After acidic elution, tryptic peptides of the A- and B-chains were obtained by accelerated digestion with trypsin in the presence of acetonitrile. Of the 20 peptides observed by MALDI-TOF MS, three were chosen for detection ( m/ z 1013.6, m/ z 1310.6 and m/ z 1728.9, which correspond to peptides 161-LEQLAGNLR-169, 150-YTFAFGGNYDR-160, and 233-SAPDPSVITLENSWGR-248, respectively). Their selection was based on several parameters such as detection sensitivity, specificity toward ricin forms and absence of isotopic overlap with unrelated peptides. To increase assay reproducibility, stable isotope-labeled peptides were incorporated during the sample preparation phase. The final assay has a limit of detection estimated at approximately 50 ng/mL ( approximately 0.8 nM) of ricin in buffer. No interference was observed when the assay was applied to ricin-spiked milk samples. In addition, several varieties of R. communis or from different geographical origins were also shown to be detectable. The present assay provides a new tool with a total analytical time of approximately 5 h, which is particularly relevant in the context of a bioterrorist incident.

Mass Spectrometry Biotyper System Identifies Enteric Bacterial Pathogens Directly from Colonies Grown on Selective Stool Culture Media

“…Targeting other pathogen-specific biomarkers has been used for EHEC identification by MALDI-TOF MS (13). Sensitive approaches that directly target functional Shigella and EHEC toxins by incorporating an immunoaffinity isolation (10,35) or an enzymatic digestion (2,20) are under investigation.…”

Ricin detection: Tracking active toxin

“…By bypassing traditional antibodies, improvements can be made in regard to reagent stability and storage life. Finally, by combining sample enrichment steps (using ricin-specific antibodies, aptamers, or sugar-conjugated materials) with enhanced detection technologies, such as surface plasmon resonance (SPR) (Blome et al, 2010;Feltis et al, 2008;Nagatsuka et al, 2013;Tran et al, 2008;Uzawa et al, 2008), polymerase chain reaction (PCR) (He et al, 2010;Lubelli et al, 2006), or mass spectrometry (MS) (Duriez et al, 2008;Kanamori-Kataoka et al, 2011), an added level of specificity and sensitivity can be achieved. Despite the many advantages associated with immunochemical methods for ricin detection, a major disadvantage exists in the inability to distinguish between biologically active and inactive ricin.…”

Analysis of lysozyme in cheese by immunocapture mass spectrometry

“…Further peak identification by the peptide mass fingerprint is not really essential for the analysis of lysozyme in cheese in routine analysis, because lysozyme could be correctly identified in all samples by its specific binding to the antibody as well as by its mass. Nevertheless, the option to verify signal identity by peptide mass fingerprint can be interesting if very similar proteins have to be distinguished, which have similar masses and epitopes [29].…”