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Single-Domain Antibodies for Intracellular Toxin Neutralization
Timothy F Czajka1, Nicholas J Mantis2,3
1Department of Biomedical Sciences, University at Albany School of Public Health, Albany, NY, USA.
Methods in Molecular Biology (Clifton, N.J.)
|February 14, 2022
Summary
Researchers developed a novel DNA-based method to neutralize ricin toxin A chain (RTA) inside cells using alpaca-derived antibodies. This approach offers a promising strategy against intracellular toxins and pathogens.
Area of Science:
- Biochemistry and Molecular Biology
- Immunology
- Biotechnology
Background:
- Ricin is a potent plant-derived toxin and a significant biothreat agent.
- The ricin toxin A chain (RTA) inhibits protein synthesis by inactivating ribosomes within the cytoplasm.
- Intracellular RTA is protected from circulating antibodies, posing a challenge for neutralization.
Purpose of the Study:
- To develop and evaluate a method for neutralizing RTA within the cytoplasm of mammalian cells.
- To utilize DNA-based delivery of single-domain antibodies (VHHs) for intracellular RTA neutralization.
Main Methods:
- Design and construction of VHH expression vectors targeting RTA's active site.
- Transient expression of VHHs in Vero cells.
- Assessment of VHH expression using enzyme-linked immunosorbent assay (ELISA) and western blotting.
- Evaluation of cellular protection through cytotoxicity studies.
Main Results:
- Successful design and expression of VHHs targeting RTA's active site within Vero cells.
- Demonstrated neutralization of RTA's cytotoxic effects in cell-based assays.
- Validation of the DNA-based VHH delivery system for intracellular toxin neutralization.
Conclusions:
- DNA-based delivery of VHHs is an effective strategy for neutralizing intracellular RTA.
- This methodology provides a versatile platform for combating ricin toxicity.
- The approach can be adapted for neutralizing other intracellular toxins and pathogens, including viruses.
