Detection of ricin and other ribosome-inactivating proteins by an immuno-polymerase chain reaction assay

Lubelli, Chiara; Chatgilialoglu, Alexandros; Bolognesi, Andrea; Strocchi, Paola; Colombatti, Marco; Stirpe, Fiorenzo

doi:10.1016/j.ab.2006.05.003

Cited by 74 publications

(56 citation statements)

References 46 publications

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“…There are two major toxins found in mature castor seeds (MOSHKIN, 1986). The first of these is ricin, a two-chain polypeptide with an LD 50 of 30 ppm, which is one of the most toxic compounds produced in nature.…”

Section: Introductionmentioning

confidence: 99%

“…The traditional methods for ricin detection use antibody-based immunoassays (LIGLER et al, 2003;RUBINA et al, 2005) and Enzyme Linked immune-sorbent assays (ELISA) (POLI et al, 1994). However, these methods require the use of animals to produce antibodies, which has been more difficult with recent animal protection policies.…”

Section: Introductionmentioning

confidence: 99%

“…The RID assay is relatively inexpensive but does require special equipment. According to Lubelli et al (2006), the LOD of IPCR was more than 1 million times lower than that of ELISA. IPCR appears to be the most sensitive method for the detection of ricin and other RIPs.…”

Section: Introductionmentioning

confidence: 99%

“…A Radial Immune Diffusion (RID) assay was adapted to facilitate screening for castor lines with reduced levels of ricin + RCA (PINKERTON et al, 1999). The RID assay uses a specific antibody against Ricinus communis lecithin incorporated into a thin layer of agar on which the antigens contained in the seed extract were placed.…”

Section: Introductionmentioning

confidence: 99%

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Development of a bioassay to quantify the ricin toxin content of castor bean (Ricinus communis L.) seeds

2012

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ABSTRACT. In this study, we developed a bioassay to quantify the ricin toxin content of castor bean seeds. Existing quantification methods do not always reflect actual toxicity of the seeds analyzed, which may present lower ricin content even though they are more toxic than seeds presenting a higher content of ricin. This is because these methods actually measure the addition of ricin RCA, which is a compound less toxic than pure ricin. We decided to use in this study, the nematode Caenorhabditis elegans, which has been widely used by the pharmaceutical industry. We tested two strains of C. elegans using different methods in 8 experiments. We examined 4 methods of extracting the ricin complex and 3 methods of exposing the nematodes. Among the nematode strains and ricin extraction methods tested, we concluded that the best strain for testing ricin toxicity was the strain called N2 and that the best method for ricin extraction was a rotating bath followed by centrifugation and exposing the nematodes in 24 well plates with a solution of nematodes extracted from the media with destilated waterexposing the nematodes in 24-well plates This method is inexpensive, quick and adequate for the selection of offspring with lower RIP content.Keywords: toxin, C. elegans, selection, Ricinus. Desenvolvimento de um bioensaio para quantificar o teor da toxina ricina em sementes de mamona (Ricinus communis L.)RESUMO. Foi desenvolvido um bioensaio para quantificar o teor de ricina nas sementes de mamona. Os métodos de quantificação existentes não refletem a toxidez real das sementes analizadas, que pode mostrar um resultado de menor teor de ricina e ainda assim ser mais toxico do que as sementes que apresentaram teores maiores. Isto ocorre por que estes métodos medem alem da ricina a RCA, que é um composto menos tóxico que a ricina pura. Foi decidido utilizar neste estudo o nematoide Caenorhabditis elegans, que tem sido amplamente utilizado na industria farmacêutica. Foram testadas duas estirpes de C.elegans usadas em diferentes métodos contando com 8 experimentos distintos. nós examinamos 4 métodos de extração de ricina e 3 métodos de exposição do nematóide à ricina. Foi concluído que a melhor estirpe para testar ricina foram as estirpes chamadas de N2 e o melhor método de extração da toxina foi o do banho maria seguido pela centrifugação e o melhor método de exposição dos nematóides foi o que conta com uma placa de 24 poços onde o nematóide suspenso em água destilada são expostos a toxina. Este é um método barato, rápido e adequado para seleção de materiais com baixo teor de RIP.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Development of a bioassay to quantify the ricin toxin content of castor bean (Ricinus communis L.) seeds

2012

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“…These advantages make them suitable for analysis of biotoxin agents and forensic samples of microscopic volumes and low concentrations. Assays based on other antibodies for ricin with higher sensitivity have been reported previously [28][29][30] . However, these assays use larger sample volumes in comparison to the droplet based microbioassay.…”

Section: Gold Only Agglutination Microbioassay With Ricin Antibodymentioning

confidence: 99%

Development and evaluation of realistic microbioassays in freely suspended droplets on a chip

Rastogi

Velev

2007

Biomicrofluidics

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A novel technique for biomolecular detection in microliter droplets floating on the surface of high density oil is presented. Each droplet was captured and manipulated dielectrophoretically and was used as a site for a microscopic bioassay based on agglutination of antibody-conjugated particles. The results were read out by the pattern of unagglomerated gold nanoparticles collected on the droplet surface. Two formats of bioassays, namely gold only agglutination and gold and latex agglutination, were investigated experimentally by varying analyte concentration, particle size and concentration, number of antigen binding sites per particle, time for incubation, and rate of particle collection on the droplet surface. The microbioassays performance was also evaluated with ricin antibodies and compared to the ricin assays in field use. It is estimated that the droplet based assays require 100ϫ smaller sample volume and are ten times more sensitive, though they require longer times to complete. The experiments were interpreted by modeling the kinetics of particle agglutination and mass transfer processes inside the droplets. The incubation time and antigen concentration values calculated by the model correlate well with the experimental results. The results could allow for development of efficient immunoassays on a chip requiring even smaller sample volumes.

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Ricin Analysis

Kalb

Barr

2014

Encyclopedia of Analytical Chemistry

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Ricin is a protein toxin produced by the castor bean plant, Ricinus communis . In vivo, the 64‐ kDa ricin protein halts protein synthesis by depurinating 28S ribosomal RNA , rendering it unable to bind to elongation factor 2. This halt in protein synthesis causes detrimental health effects and can lead to death upon exposure to ricin. Ricin is highly toxic and has the potential for high availability; therefore, it is considered a likely agent for bioterrorism. Hence, sensitive and selective analytical methods are desired to analyze for ricin in food, environmental, and clinical samples. This article reviews various published methods to detect ricin. These methods are numerous but can be loosely grouped into animal assays, detection of ricin DNA , immunoassays, mass‐spectrometry‐based amino acid composition methods, and detection of ricin enzymatic activity. These various methods have different purposes. Some methods function as rapid screens with moderate false positive and false negative rates, whereas others serve as confirmatory assays. In this article, the methods are explained and compared, with discussion on the sensitivity and selectivity of the methods.

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Detection of ricin and other ribosome-inactivating proteins by an immuno-polymerase chain reaction assay

Cited by 74 publications

References 46 publications

Development of a bioassay to quantify the ricin toxin content of castor bean (Ricinus communis L.) seeds

Development of a bioassay to quantify the ricin toxin content of castor bean (Ricinus communis L.) seeds

Development and evaluation of realistic microbioassays in freely suspended droplets on a chip

Ricin Analysis

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