Titration of Botulinum Toxins for Lethal Toxicity by Intravenous Injection Into Mice

Kondo, Hisashi; Shimizu, Tomoko; Kubonoya, Masaru; Izumi, Nanae; Takahashi, Motohide; Sakaguchi, Genji

doi:10.7883/yoken1952.37.131

Cited by 53 publications

(22 citation statements)

References 2 publications

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“…BoNTs were purified principally according to the method of Kurazono et al (22). Toxicity was assayed by the timeto-death method by intravenous injection into mice (23). Samples were also titrated by intraperitoneal injection into mice with serial 2-fold dilutions to obtain a mean 50% lethal dose (LD 50 ) by the calculations of Reed and Muench (24).…”

Section: Methodsmentioning

confidence: 99%

Binding of Clostridium botulinum Type C and D Neurotoxins to Ganglioside and Phospholipid

Tsukamoto¹,

Kohda²,

Mukamoto³

et al. 2005

Journal of Biological Chemistry

122

117

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Clostridium botulinum neurotoxins (BoNTs) act on nerve endings to block acetylcholine release. Their potency is due to their enzymatic activity and selective high affinity binding to neurons. Although there are many pieces of data available on the receptor for BoNT, little attempt has been made to characterize the receptors for BoNT/C and BoNT/D. For this purpose, we prepared the recombinant carboxyl-terminal domain of the heavy chain (H C ) and then examined its binding capability to rat brain synaptosomes treated with enzymes and heating. Synaptosomes treated with proteinase K or heating retained binding capability to both H C /C and H C /D, suggesting that a proteinaceous substance does not constitute the receptor component. We next performed a thin layer chromatography overlay assay of H C with a lipid extract of synaptosomes. Under physiological or higher ionic strengths, H C /C bound to gangliosides GD1b and GT1b. These data are in accord with results showing that neuraminidase and endoglycoceramidase treatment decreased H C /C binding to synaptosomes. On the other hand, H C /D interacted with phosphatidylethanolamine but not with any ganglioside. Using cerebellar granule cells obtained from GM3 synthase knock-out mice, we found that BoNT/C did not elicit a toxic effect but that BoNT/D still inhibited glutamate release to the same extent as in granule cells from wild type mice. These observations suggested that BoNT/C recognized GD1b and GT1b as functional receptors, whereas BoNT/D induced toxicity in a ganglioside-independent manner, possibly through binding to phosphatidylethanolamine. Our results provide novel insights into the receptor for clostridial neurotoxin.

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Section: Methodsmentioning

confidence: 99%

Binding of Clostridium botulinum Type C and D Neurotoxins to Ganglioside and Phospholipid

Tsukamoto¹,

Kohda²,

Mukamoto³

et al. 2005

Journal of Biological Chemistry

122

117

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show abstract

“…Toxicity was determined by the time-to-death method in mice (9). Protein contents were determined by the method of Lowry et al (10).…”

Section: Methodsmentioning

confidence: 99%

Comparative Studies on Clostridium Botulinum Type a Strains Associated With Infant Botulism in Japan and in California, Usa

Tabita

Sakaguchi

Kozaki

et al. 1990

JJMSB

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“…The radioiodinated protein was separated from reactants on a Sephadex G-50 column. Protein concentration was determined by the methods of Lowry et al (1951) or Bradford (1976), and toxicity was bioassayed by the methods of Hardegree (1965) and Kondo et al (1984). Residual toxicity of iodinated preparations was 65 to 85 percent.…”

Section: B Methodsmentioning

confidence: 99%

“…The in vivo potency of clostridial toxins was determined by the methods of Hardegree (1965) and Kondo et al (1984). In vitro toxicity was bioassayed on the mouse phrenic nerve-hemidiaphragm preparation as previously described (Simpson, 1980).…”

Section: B Methodsmentioning

confidence: 99%

A Core Facility for the Study of Neurotoxins of Biological Origin

Simpson¹

1992

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Titration of Botulinum Toxins for Lethal Toxicity by Intravenous Injection Into Mice

Cited by 53 publications

References 2 publications

Binding of Clostridium botulinum Type C and D Neurotoxins to Ganglioside and Phospholipid

Binding of Clostridium botulinum Type C and D Neurotoxins to Ganglioside and Phospholipid

Comparative Studies on Clostridium Botulinum Type a Strains Associated With Infant Botulism in Japan and in California, Usa

A Core Facility for the Study of Neurotoxins of Biological Origin

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