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.