Abstract
Abrin, one of the most highly potent toxins in the world, is derived from the plant, Abrus precatorius. Because of its high toxicity, it poses potential bioterror risks. Therefore, a need exists for new reagents and technologies that would be able to rapidly detect abrin contamination as well as lead to new therapeutics. We report here a group of abrin-specific monoclonal antibodies (mAbs) that recognize abrin A-chain, intact A–B chain toxin, and agglutinin by Western blot. Additionally, these mAbs were evaluated for their ability to serve as capture antibodies for a sandwich (capture) ELISA. All possible capture–detector pairs were evaluated and the best antibody pair identified and optimized for a capture ELISA. The capture ELISA based on this capture–detector mAb pair had a limit of detection (L.O.D) of ≈1 ng/mL measured using three independent experiments. The assay did not reveal any false positives with extracts containing other potential ribosome-inactivating proteins (RIPs). Thus, this new capture ELISA uses mAbs for both capture and detection; has no cross-reactivity against other plant RIPs; and has a sensitivity comparable to other reported capture ELISAs using polyclonal antibodies as either capture or detector.
Highlights
Abrin and ricin are both members of the Type II family of ribosome-inactivating proteins (RIP)that inhibit eukaryotic protein synthesis leading to apoptosis and cell death
Anti-abrin monoclonal antibodies (mAbs) from ten hybridoma cell lines were grown as previously described [24] and purified from the ascites fluid by affinity chromatography on Protein-G Sepharose
LS13ABx as as aa detector detector are are shown shown in in a using capture with with biotin-labeled biotin-labeled mAb a representative ELISA curve in Figure 3 representing one independent experiment consisting of three replicates
Summary
Abrin and ricin are both members of the Type II family of ribosome-inactivating proteins (RIP). That inhibit eukaryotic protein synthesis leading to apoptosis and cell death. Abrin like ricin is considered a Select Agent due to its potential use in bio warfare. Abrin is a heterodimeric protein toxin consisting of an A-chain and a B-chain linked together with a single disulfide. The catalytic active A-chain is an N-glycosidase that deadenylates the rat 28S ribosomal. RNA at position 4324 [1] thereby preventing ribosomes from binding to elongation factor (EF) 1 and 2 leading to inhibition of protein synthesis and eventual cell death [2,3,4,5,6,7]. Abrin B-chain is a lectin that binds to cell surface carbohydrate receptors facilitating receptor-mediated endocytosis of the
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