Comparison of the primary rat spinal cord cell (RSC) assay and the mouse bioassay for botulinum neurotoxin type A potency determination

Pellett, Sabine; Tepp, William H.; Toth, Stephen I.; Johnson, Eric A.

doi:10.1016/j.vascn.2010.01.003

Cited by 57 publications

(69 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Only an assay that considers these four sequential steps can allow an accurate determination of BoNT potency. As supported here, neuronal cell-based (NCB) assays, which meet all of these requirements, are excellent models for determination of BoNT potency (28,29,(47)(48)(49)(50)(51)(52)(53). However, correlation to the MBA still has to be confirmed for most of these assays, and the presented data suggest that this correlation should be determined for each BoNT subtype and formulation.…”

Section: Discussionmentioning

confidence: 80%

“…Primary mouse spinal cord (MSC) cells (31,41), primary rat spinal cord (RSC) cells, and human induced pluripotent stem cell (hiPSC)-derived neurons from two different sources (iCell Neurons and HIP neural stem cell-derived neurons) were treated with serial dilutions of the five BoNT/A subtypes for 48 h. The iCell Neurons comprise a mixture of primarily glutamatergic neurons and GABAergic neurons similar to that seen with forebrain cells, and the differentiated HIP neural stem cells primarily comprise a mixture of cholinergic, GABAergic, and glutamatergic neurons and were directed to enrich the motor neuron population by addition of retinoic acid and purmorphamine. EC 50 s for BoNT/A1 in RSC cells and iCell Neurons have previously been documented to be between 0.2 and 0.3 U or around 0.25 pM (28)(29)(30).…”

Section: Resultsmentioning

confidence: 98%

“…Primary rat spinal cord (RSC) cells and human induced pluripotent stem cell (hiPSC)-derived neurons (iCell Neurons; Cellular Dynamics International, Madison, WI) were prepared and maintained in serum-free medium as previously described (28)(29)(30). Primary mouse spinal cord (MSC) cells were prepared essentially as previously described (31).…”

Section: Methodsmentioning

confidence: 99%

“…For all neuronal cell-based (NCB) assays, the neuronal cells were exposed to the indicated concentrations of BoNT/A subtypes in 50 l of the respective culture medium for 48 h. All samples were tested in a minimum of triplicate experiments, and a negative control without BoNT was included. After the specified exposure time, the extracellular solution containing BoNT was removed and cells were lysed in 1ϫ lithium dodecyl sulfate (LDS) sample buffer (Invitrogen) and analyzed by Western immunoblotting for SNAP-25 cleavage as previously described (28)(29)(30). Cleaved and uncleaved SNAP-25 bands were quantified by densitometry using a Foto/Analyst FX system and TotalLab Quant software (Fotodyne).…”

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

et al. 2013

Self Cite

View full text Add to dashboard Cite

Botulinum neurotoxins (BoNTs) are synthesized by Clostridium botulinum and exist as seven immunologically distinct serotypes designated A through G. For most serotypes, several subtypes have now been described based on nominal differences in the amino acid sequences. BoNT/A1 is the most well-characterized subtype of the BoNT/A serotype, and many of its properties, including its potency, its prevalence as a food poison, and its utility as a pharmaceutical, have been thoroughly studied. In contrast, much remains unknown of the other BoNT/A subtypes. In this study, BoNT/A subtype 1 (BoNT/A1) to BoNT/A5 were characterized utilizing a mouse bioassay, an in vitro cleavage assay, and several neuronal cell-based assays. The data indicate that BoNT/A1 to -5 have distinct in vitro and in vivo toxicological properties and that, unlike those for BoNT/A1, the neuronal and mouse results for BoNT/A2 to -5 do not correlate with their enzymatic activity. These results indicate that BoNT/A1 to -5 have distinct characteristics, which are of importance for a greater understanding of botulism and for pharmaceutical applications.

show abstract

Section: Discussionmentioning

confidence: 80%

Section: Resultsmentioning

confidence: 98%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

et al. 2013

Self Cite

View full text Add to dashboard Cite

show abstract

“…To determine whether immunization with HcmR4 induces neutralizing antibodies to Hc, we used the rat spinal cord cell (RSC) assay (34,35). The RSC assay has been validated against the mouse lethality assay for assessing BoNT/A neutralizing antibodies in sera (34).…”

Section: Downloaded Frommentioning

confidence: 99%

Rapid Immune Responses to a Botulinum Neurotoxin Hc Subunit Vaccine through In Vivo Targeting to Antigen-Presenting Cells

et al. 2011

Self Cite

View full text Add to dashboard Cite

Botulinum neurotoxins A, B, C, E, and F preferentially enter cultured human motor neurons compared to other cultured human neuronal populations

2019

Self Cite

View full text Add to dashboard Cite

Human‐induced pluripotent stem cell (hiPSC)‐derived neurons can be exquisitely sensitive to botulinum neurotoxins (BoNTs), exceeding sensitivity of the traditionally used mouse bioassay. In this report, four defined hiPSC‐derived neuronal populations including primarily GABAergic, glutamatergic, dopaminergic, and motor neurons were examined for BoNT/A, B, C, D, E, and F sensitivity. The data indicate that sensitivity varies markedly for the BoNTs tested. Motor neurons are significantly more sensitive than other neuron types for all BoNTs except BoNT/D. Examination of SNARE protein levels and BoNT‐specific cell surface protein receptors reveals few differences between the cell types except greater expression levels of the receptor protein SV2C and synapsin‐IIa in motor neurons. This indicates that differential toxicity of BoNTs for motor neurons compared to other neuronal cell types involves multiple mechanisms.

show abstract

Comparison of the primary rat spinal cord cell (RSC) assay and the mouse bioassay for botulinum neurotoxin type A potency determination

Cited by 57 publications

References 32 publications

Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

Characterization of Botulinum Neurotoxin A Subtypes 1 Through 5 by Investigation of Activities in Mice, in Neuronal Cell Cultures, and In Vitro

Rapid Immune Responses to a Botulinum Neurotoxin Hc Subunit Vaccine through In Vivo Targeting to Antigen-Presenting Cells

Botulinum neurotoxins A, B, C, E, and F preferentially enter cultured human motor neurons compared to other cultured human neuronal populations

Contact Info

Product

Resources

About