Immunosorbent analysis of ricin contamination in milk using colorimetric, chemiluminescent and electrochemiluminescent detection

Brandon, David L.; Korn, Anna M.; Yang, Lily L.

doi:10.1080/09540105.2012.753515

Cited by 10 publications

(11 citation statements)

References 33 publications

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“…The differences between the types of milk and between buffer and the average of the milk data were not significant (p > 0.1) for either k a or k d . These results are consistent with our report that mAb 1797 was a highly effective capture antibody in three immunosorbent assay formats for ricin in the milk matrix (Brandon et al 2014).…”

Section: Biosensor Studies Of Ricin Monoclonal Antibodies 1755supporting

confidence: 95%

Antibody Interactions withRicinus communisAgglutinins Studied by Biolayer Interferometry

Brandon¹,

Adams²,

Yang³

et al. 2014

Analytical Letters

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Two related agglutinins are present in the seeds of Ricinus communis (castor): ricin, a dichain ribosome-inactivating protein and Ricinus communis agglutinin-1, a much less toxic tetrameric hemagglutinin. The immunochemical analysis of these agglutinins is of special interest because ricin toxicity has resulted in both accidental and intentional poisonings, while it has also provided a potential cancer chemotherapeutic in the form of an immunoconjugate. We previously characterized a panel of monoclonal antibodies (mAbs) for the analysis of potential contamination with ricin in several food matrices. In this study, an optical sensing technique, biolayer interferometry (BLI), was used to study the binding of two mAbs to the agglutinins. MAbs were immobilized on sensors with amine-reactive, Fc-binding, and streptavidin-coated tips to study the interactions with the agglutinins and with ricin A-and B-chains in solution. The kinetically determined equilibrium dissociation constants generally agreed with the relative binding observed in ELISA, although binding was less predictable for the isolated ricin chains. BLI analysis of kinetic constants for mAb 1797 was not affected by nonfat milk (0.5% by volume). BLI provides a useful method to characterize the binding of antibodies, with the potential for immunodiagnostic applications in food matrices.

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Section: Biosensor Studies Of Ricin Monoclonal Antibodies 1755supporting

confidence: 95%

Antibody Interactions withRicinus communisAgglutinins Studied by Biolayer Interferometry

Brandon¹,

Adams²,

Yang³

et al. 2014

Analytical Letters

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“…All mAbs had been elicited in mice by immunization protocols beginning with inoculation of ricin A chain [10][11][12][13], and all mAbs, except mAb 1797, bound to wells coated with isolated A chain in direct ELISA. We had previously identified many apparently A-chain-specific mAbs that did not bind to solid-phase isolated chain, perhaps because of loss of conformational epitopes after the disulfides linking the A and B chains were disrupted or because of structural alteration upon adsorption onto the plastic ELISA wells [18].…”

Section: Resultsmentioning

confidence: 99%

“…Mouse monoclonal antibodies were prepared in our laboratory to develop ELISA, immuno-PCR, and electrochemical immunosorbent methods that could detect potential ricin contamination of milk, liquid egg, and ground beef, and for toxicokinetic studies [6,[10][11][12][13]. They are denoted by the serial number of the hybridoma clone from which they were obtained, as shown in Table 1.…”

Section: Antibodies and Elisa Reagentsmentioning

confidence: 99%

“…Previous research identified means to distinguish gene expression of ricin and RCA [9], but this approach is not applicable for protein levels. The development of monoclonal antibodies (mAbs) to determine castor agglutinins was previously reported and applied to food analyses [10][11][12][13] and ricin toxicokinetics [6]. Discrimination between ricin and RCA-1 is not critical for screening of food samples, as neither ricin nor RCA should be present.…”

Section: Introductionmentioning

confidence: 99%

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Analysis of Castor by ELISAs that Distinguish Ricin and Ricinus communis agglutinin (RCA)

Brandon

McKeon

Patfield

et al. 2015

J Americ Oil Chem Soc

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To facilitate the analysis of castor (Ricinus communis L.) seed fractions and germplasm for ricin content, we investigated the use of enzyme‐linked immunosorbent assay (ELISA) methods to differentiate between ricin toxin and the related Ricinus communis agglutinin (RCA). Both proteins are based on a heterodimeric AB structure, with a common A chain. RCA comprises two dimers of A and B chains. Both proteins are found in the meal remaining after castor oil extraction and impede the commercial production of castor seed in the USA. We identified pairs of monoclonal antibodies (mAbs) that could distinguish between the structurally related proteins that share a common A chain. Antibody specificity was determined by ELISA and checked by immunoblotting. We found that mAb–mAb pairs afforded quantification of each castor protein, and that a mAb paired with a commercial polyclonal antibody provided detection of both with comparable sensitivity.

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“…A cadeia A é uma potente ribotoxina, que inibe a síntese de proteínas em células; enquanto a cadeia B é uma lectina, que se liga a resíduos de galactose na membrana celular. A concentração letal da ricina está estimada em 2 mg para um adulto quando há exposição entérica (Godoy et al, 2009, Brandon et al, 2014. Devido ao risco potencial desta toxina, fica clara a necessidade de separa-la da lipase de mamona de forma que esta lipase possa ser aplicada de maneira mais generalizada e sem riscos.…”

Section: Introductionunclassified

EXTRAÇÃO DE LIPASE DE MAMONA (Ricinus communis L.) EM FASE AQUOSA

SILVA¹,

KOPP²,

GIORDANO³

2015

Anais Do XX Congresso Brasileiro De Engenharia Química

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RESUMO -Endospermas de mamona (Ricinus communis L.) processados contendo atividade lipolítica têm sido utilizados com sucesso em alguns bioprocessos. Porém, a toxina ricina presente na mamona limita a aplicação deste biocatalisador, sendo necessário separá-la da lipase. A primeira etapa deste processo é a solubilização do biocatalisador em fase aquosa. Neste contexto, foi avaliada a extração da lipase de mamona em diferentes tampões e pHs. O extrato bruto de mamona foi incubado em diversos tampões por 12h a 25°C; seguido de centrifugação para separação do endosperma. Os melhores resultados nesta etapa foram obtidos com tampão fosfato 0.05 mol.L -1 pH 7.0, carbonato-bicarbonato 0.05 mol.L -1 pH 9.0 e 10.0, onde mais de 70% da atividade lipolítica foi extraída para a fase líquida ao final do processo. INTRODUÇÃOProcessos enzimáticos, quando comparados com processos catalisados quimicamente, apresentam como principais vantagens a redução do consumo energético do processo, uma vez que enzimas atuam em condições suaves de temperatura e pressão, apresentando alta seletividade e especificidade (Narwal, Gupta 2013). Enzimas são capazes de catalisar reações químicas sem a formação de subprodutos indesejáveis e sem as custosas etapas de proteção e desproteção de grupos reativos necessárias na maioria dos processos onde catalisadores químicos são empregados (Leung et al., 2010).Atualmente, inúmeras enzimas são empregas em processos industriais, como proteases, celulases e lipases. Dentre estas, lipases (glicerol éster hidrolases, E.C 3.1.1.3) são especialmente interessantes devido sua vasta aplicabilidade -desde a indústria alimentícia, passando pela química fina chegando até a produção de biocombustíveis -fazendo com que sejam alvo de extensa pesquisa científica (Vescovi 2012).Lipases naturalmente catalisam a hidrólise de óleos e gorduras quando em meio aquoso, de forma parcial ou total. Porém, também são capazes de realizar reações de esterificação, transesterificação (alcoólise) e interesteríficação de lipídeos quando em meio orgânico (Atabani et al., 2012;Adlercreutz 2013). Embora lipases mais utilizadas industrialmente sejam obtidas a partir de fontes microbianas, lipases podem ser obtidas de diferentes fontes, como células animais e vegetais (Pierozan et al., 2009). Enzimas vegetais são novidade em termo de fontes enzimáticas; porém possuem vantagens como baixo custo da matéria-prima, aplicação na indústria farmacêutica e alimentícia, fácil aceitação no mercado devido sua Área temática: Processos Biotecnológicos 1

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Immunosorbent analysis of ricin contamination in milk using colorimetric, chemiluminescent and electrochemiluminescent detection

Cited by 10 publications

References 33 publications

Antibody Interactions withRicinus communisAgglutinins Studied by Biolayer Interferometry

Antibody Interactions withRicinus communisAgglutinins Studied by Biolayer Interferometry

Analysis of Castor by ELISAs that Distinguish Ricin and Ricinus communis agglutinin (RCA)

EXTRAÇÃO DE LIPASE DE MAMONA (Ricinus communis L.) EM FASE AQUOSA

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