Development of an ELISA microarray platform for the sensitive and quantitative detection of botulinum neurotoxins

Zhang, Yuchen; Lou, Jianlong; Marks, James D.; Jenko, Kathryn L.; Varnum, Susan M.

doi:10.1016/j.toxicon.2012.07.117

Cited by 1 publication

(1 citation statement)

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“…For instance, the conventional ELISA test is less sensitive than the MBA and cross-reactivity between serotypes can be problematic (Wilder-Kofie et al, 2011). Even a more recently described high-affinity monoclonal antibody capture ELISA was seven times less sensitive than the modified FDC assay (Zhang et al, 2012) while a BoTest immunoprecipitation method was no more sensitive (LoD 0.1 pM,~3 LD 50 per ml serum) than the MBA (Dunning et al, 2012). Additionally, sensitive (LoD 0.01 LD 50 ml -1 ) direct endopeptidase tests (Jones et al, 2008) are adversely affected when partially or undiluted serum is used (R. G. A.…”

Section: Discussionmentioning

confidence: 99%

Use of a new functional dual coating (FDC) assay to measure low toxin levels in serum and food samples following an outbreak of human botulism

Jones

Marks

2013

Journal of Medical Microbiology

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Clostridium botulinum type A toxin is the most prevalent cause of naturally occurring outbreaks of human botulism in the world. The active dichain neurotoxin molecule is composed of a heavy chain (H-chain) of~100 kDa with the carboxy-terminal end consisting of a receptor-binding (H C ) domain, while the amino-terminal (H N ) domain is linked by a critical disulfide bond to a light chain (L-chain) of~50 kDa. Although the mouse bioassay (MBA) is traditionally used to confirm the presence of toxin in serum or food, its sensitivity is insufficient to detect low toxin levels in approximately 30 to 60 % of botulism patients. A novel FDC (functional dual coating) microtitre plate immuno-biochemical assay, which quantifies botulinum toxicity by measuring the H C domain linked with L-chain endopeptidase activity, was modified to allow human serum (lysed or unlysed) to be tested without interference from the matrix, with toxin detection down to 0.03 mouse LD 50 per ml serum or 0.13 pg ml -1 using just 100 ml of clinical samples. The assay was specific for type A toxin and could additionally be applied to whole blood and food samples. Low levels of 1 to 2 mouse LD 50 per ml serum of type A toxin were quantified for the first time using the modified FDC assay in two severely intoxicated UK patients who required mechanical ventilation and antitoxin. Toxin levels in recovered food sample extracts were also detected and one MBAnegative sample was found to contain 0.32 LD 50 per ml extract. The FDC assay provides a real alternative for public health laboratories to unambiguously confirm all cases of type A botulism and, due to its sensitivity, a promising new tool in toxin pharmacokinetic studies.

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Section: Discussionmentioning

confidence: 99%