Douglas C. Stafford

Development of an Avian Antitoxin to Type A Botulinum Neurotoxin

Most commercially available antitoxins and antivenoms are raised in horses and purified by bulk fractionation techniques. These preparations frequently elicit deleterious side effects that compromise their efficacy and the treatment of intoxication or envenomation. Alternatively, the use of hyperimmune immunoglobulin from the egg yolks of laying hens has shown considerable promise as a cost-effective and potentially safer source of antitoxins and antivenoms. To investigate the utility of this system in the development of a botulism antitoxin, laying hens were immunized with purified 150 kDa type A botulinum neurotoxin detoxified with formaldehyde. Antibodies present in egg yolk were then isolated by polyethylene glycol (PEG) fractionation and further purified by affinity chromatography using immobilized toxoid. These antibodies exhibited a high titer of toxoid reactivity as measured by enzyme immunoassay. A mouse lethality/protection assay demonstrated the efficacy of these antibodies in neutralizing type A botulinum neurotoxin, where PEG-fractionated and affinity-purified antibodies neutralized 38 and 340 I.U. of type A botulinum neurotoxin per milligram of protein, respectively. Thus, the avian antibody preparations, even if only PEG-fractionated, are significantly more potent (neutralization titer per mg protein) than conventional equine botulism antiserum preparations. In addition to their superior potency, hyperimmune avian antibodies, whether PEG-fractionated or affinity-purified, are expected to significantly increase antitoxin safety.