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Detecting and preventing reversion to toxicity for a formaldehyde-treated C. difficile toxin B mutant
Bei Wang1, Su Wang1, Richard R Rustandi1
1Merck Research Laboratories, 770 Sumneytown Pike, West Point, PA 19486, USA.
Vaccine
|June 22, 2014
Summary
Detoxifying Clostridium difficile toxin B (TcdB) with formaldehyde is effective, but liquid storage causes toxicity reversion. Lyophilization of formaldehyde-treated TcdB prevents this reversion, maintaining stability and immunogenicity.
Area of Science:
- Microbiology
- Biochemistry
- Immunology
Background:
- Clostridium difficile toxin B (TcdB) is highly toxic.
- A five-point mutation TcdB mutant (mTcdB) retains residual toxicity.
- Formaldehyde treatment detoxifies mTcdB, but liquid storage leads to toxicity reversion.
Purpose of the Study:
- To investigate methods for preventing toxicity reversion in formaldehyde-treated mTcdB.
- To assess the stability and immunogenicity of lyophilized mTcdB.
Main Methods:
- Treatment of mTcdB with formaldehyde.
- Storage of liquid and lyophilized mTcdB formulations at 37°C.
- Toxicity assays in cell cultures and mice.
- Biochemical analyses including SDS-PAGE and anion exchange chromatography.
- Immunogenicity testing in mice.
Main Results:
- Liquid storage of formaldehyde-treated mTcdB resulted in toxicity reversion, temperature-dependent.
- Lyophilization of formaldehyde-treated mTcdB prevented toxicity reversion, even after 6 months at 37°C.
- Lyophilized mTcdB remained stable and immunogenic post-storage.
Conclusions:
- Lyophilization is a superior method for stabilizing formaldehyde-detoxified mTcdB, preventing toxicity reversion.
- Formaldehyde-treated, lyophilized mTcdB is a stable and potentially viable immunogen.
