Summary
125I-labelled tetanus toxin and 125I-labelled botulinum A neurotoxin are known to be specifically bound to brain synaptosomes. In order to discriminate between active toxin and inactive admixtures present in the starting material or arising during isodination, synaptosome columns were prepared using bromacetylcellulose and/or kieselgur (Celite®) as carriers. Both types of columns adsorb the toxins from low ionic strength medium and release them if the pH and ionic strength are raised.
Botulinum toxin was eluted with lower ionic strength than tetanus toxin, and could be freed from nontoxic admixtures.
Analysis by affinity chromatography disclosed partially toxoided tetanus toxin in both labelled and unlabelled toxin samples. High concentrations of formaldehyde (0.5%) destroyed both toxicity and affinity to the synaptosomes of tetanus toxin. Low concentrations of formaldehyde (0.05%) yielded a derivative of low toxicity which was still, however less firmly, bound to synaptosomes.
Tetanus and botulinum toxin differ by their acceptors. Whereas unlabelled botulinum toxin is unable to compete with labelled tetanus toxin, unlabelled tetanus toxin slightly competes with botulinum toxin. Both labelled toxins display anomalous binding behaviour in that they cannot be displaced completely even with a large excess of unlabelled toxin.
Similar content being viewed by others
References
Habermann, E.: Distribution of 125I-tetanus toxin and 125I-toxoid in rats with local tetanus, as influenced by antitoxin. Naunyn-Schmiedeberg's Arch. Pharmacol. 272, 75–88 (1972)
Habermann, E.: Discrimination between binding to CNS, toxicity and immunreactivity of derivatives of tetanus toxin. Med. Microbiol. Immunol. 159, 89–100 (1973a)
Habermann, E.: Interaction of labeled tetanus toxin and toxoid with substructures of rat brain and spinal cord in vitro. Naunyn-Schmiedeberg's Arch. Pharmacol. 276, 341–359 (1973b)
Habermann, E.: 125I-labeled neurotoxin from clostridium botulinum A: Preparation, binding to synaptosomes and ascent to the spinal cord. Naunyn-Schmiedeberg's Arch. Pharmacol. 281, 47–56 (1974)
Habermann, E., Dimpfel, W.: Distribution of 125I-tetanus toxin and 125I-toxoid in rats with generalized tetanus, as influenced by antitoxin. Naunyn-Schmiedeberg's Arch. Pharmacol. 276, 327–340 (1973)
Habermann, E., Dimpfel, W., Räker, K. O.: Interaction of labelled tetanus toxin with substructures of rat spinal cord in vivo. Naunyn-Schmiedeberg's Arch. Pharmacol. 276, 361–373 (1973)
Habermann, E., Heller, I.: Direct evidence for the specific fixation of Cl. botulinum A neurotoxin to brain matter. Naunyn-Schmiedeberg's Arch. Pharmacol. 287, 97–106 (1975)
Hirokawa, N., Kitamura, M.: Localization of radioactive 125I-labeled botulinus toxin at the neuromuscular junction of mouse diaphragm. Naunyn-Schmiedeberg's Arch. Pharmacol. 287, 107–110 (1975)
Kryzhanovskyi, G. N.: The mechanism of action of tetanus toxin: Effect on synaptic processes and some particular features of toxin binding by the nervous tissue. Naunyn-Schmiedeberg's Arch. Pharmacol. 276, 247–270 (1973)
Lowry, O. H., Rosebrough, N. J., Farr, A. L., Randall, R. J.: Protein measurements with the Folin phenol reagent. J. biol. Chem. 193, 265–275 (1951)
Mellanby, J., Van Heyningen, W. E., Whittaker, V. P.: Fixation of tetanus toxin by subcellular fractions of brain. J. Neurochem. 12, 77–79 (1964)
Prévot, A. R.: Biologie des maladies dues aux anaerobies. Paris: Flammarion 1955
Robbins, J. B., Haimovich, J., Sela, M.: Purification of antibodies with immunoadsorbents prepared using bromacetyl cellulose. Immunochemistry 4, 11–22 (1967)
Van Heyningen, W. E.: On the similarity of tetanus and cholera toxins. Naunyn-Schmiedeberg's Arch. Pharmacol. 276, 289–295 (1973)
Whittaker, V. P.: The synaptosomes. In: Handbook of neurochemistry, vol. II (A. Lajtha, ed.), pp. 327–364. New York-London: Plenum Press 1969
Wiegand, H., Erdmann, G., Wellhöner, H. H.: 125I-labelled botulinum A neurotoxin: Pharmacokinetics in cats after intramuscular injection. Naunyn-Schmiedeberg's Arch. Pharmacol. (in press, 1976)
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Habermann, E. Affinity chromatography of tetanus toxin, tetanus toxoid, and botulinum a toxin on synaptosomes, and differentiation of their acceptors. Naunyn-Schmiedeberg's Arch. Pharmacol. 293, 1–9 (1976). https://doi.org/10.1007/BF00498864
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00498864