Antibodies to toxins

The crosslinking process must minimize polymerization of either the antibody or the toxin. Low molecular weight, 1 1 or 1 2 conjugates of antibody-to-toxin are best. 

Several studies have described the production of antibodies to toxins, that are capable of detecting very small quantities of contaminants, such as aflatoxins in grain and nut products [37-39] and shellfish poisoning toxins [40,41]. 

The best method of preparing individual toxins from the crude venom is by affinity immunochromatography utilizing monoclonal antibody or in one instance, polyclonal antibody (4—6). Monoclonal antibodies to both the fishing and mesentery... 

The crosslinker used to form the bond between the antibody and toxin must be able to survive in vivo and not be cleaved by enzymatic or reductive means before reaching the targeted cells. 

Figure 21.5 SPDP can be used to modify both an antibody and a toxin molecule for conjugation purposes. In this case, the antibody is thiolated to contain a sulfhydryl group by modification with SPDP followed by reduction with DTT. A toxin molecule is then activated with SPDP and reacted with the thiolated antibody to effect the final conjugate through a disulfide bond.

Another way of utilizing SPDP is to again activate the antibody to create the pyridyl disulfide derivative, but this time thiolate the toxin component using 2-iminothiolane (Chapter 1,... 

Figure 21.7 An intact A-B subunit toxin molecule may be activated with 2-iminothiolane with good retention of cytotoxic activity. The thiolated toxin then may be conjugated with SPDP-activated antibody to generate the immunotoxin conjugate through a disulfide bond.

Dissolve the antibody to be conjugated in 0.1 M sodium phosphate, 0.15 M NaCl, pH 7.5, at a concentration lOmg/ml. Note some protocols use the borate buffer system described in step A. Use 2.5 mg of antibody per mg of toxin to be conjugated. 

Figure 21.10 Cystamine may be used to make immunotoxin conjugates by a disulfide interchange reaction. Modification of antibody molecules using an EDC-mediated reaction creates a sulfhydryl-reactive derivative. A-chain toxin subunits containing a free thiol can be coupled to the cystamine-modified antibody to form disulfide crosslinks.

The antibody preparations could be administered unaltered or (more commonly) after their conjugation to radioisotopes or toxins. Binding of unaltered monoclonal antibodies to a tumour surface alone should facilitate increased destruction of tumour cells (Figure 13.4). This approach, however, has yielded disappointing results, as the monoclonal antibody preparations used to date have been murine in origin. The Fc region of such mouse antibodies is a very poor activator of human immune function. Technical advances, allowing the production of human/humanized monoclonals (see later) may render this therapeutic approach more attractive in the future. 

Immunization Administration either of a non-toxic antigen to confer active immunity or antibody to confer passive immunity to a person or animal in order to render them insusceptible to the toxic effects of a pathogen or toxin. 

Treatment — There is no specific treatment for C. perfringens toxins, even though the organism is susceptible to penicillin, which is the drug of choice for a naturally acquired infection. Recent laboratory data indicate that clindamycin and rifampin may suppress toxin formation. Available polyvalent antitoxins contain antibodies to several toxins that have been used in treatment, but not enough data exist to prove efficacy.3... 

A variant of the original immunotoxin approach is the so-called immunocytokines. In these constructs the antibody targeting moiety is maintained, but the toxin as the effector molecule is replaced by a cytokine. In contrast to toxins, cytokines are often proteins endogenously produced in man. If both the antibody and cytokine are of human origin, then no foreign proteins are introduced which could provoke an antibody response from the host immune system when the drug targeting preparation is clinically applied. 

Nl. Nagata, S., Tsutsumi, T., Yoshida, R, and Ueno, Y, A new type sandwich immunoassay for microcystin Production of monoclonal antibodies specific to the immune complex formed by microcystin and an anti-microcystin monoclonal antibody. Natural Toxins 1, 49-55 (1999). 

The minimum antibody binding domain, Fv, has also been linked to toxin to produce a more compact, recombinant immuno-toxin. The recombinant F -toxin appears to be more stable and exhibits less liver accumulation [27,28]. The immunogenicity of Fv-toxin as a consequence of the highly antigenic determinants of bacterial toxin remains a barrier to development. 

Gilliland, D.G., Z. Steplewski, R.J. Collier, K.F Mitchell, T.H. Chang, and H. Koprowski, Antibody-directed cytotoxic agents use of monoclonal antibody to direct the action of toxin A chains to colorectal carcinoma cells. Proc Natl Acad Sci USA, 1980. 77(8) 4539-43. 

Scientists have found a way to insert into a potato a gene that generates the protein the cholera bacterium uses to bind its toxin to cells in the gut. A person who eats this potato will form antibodies to the protein, antibodies that would recognize and destroy the binding protein should cholera bacteria be encountered. Based on animal studies, we can say that one such cooked potato a week for a month, along with periodic boosters, should do the job. It would be especially appropriate if they served such potatoes in the John Snow pub. Actually,... 

---