Detoxification of diphtheria and tetanus toxin with formaldehyde. Detection of protein conjugates.

In a model system of purified diphtheria and tetanus toxins it was shown that conjugates between the two proteins are formed during detoxification with formaldehyde. Detoxification mixtures were fractionated by HPLC. Two protein conjugates with different molecular weights were detected and quantified by capture ELISA assay. In vivo the existence of the largest diphtheria-tetanus toxoid conjugate was demonstrated by its antibody response in mice vaccinated with a calcium phosphate adjuvated column fraction of detoxification mixture. To eliminate the risk of cross-linking foreign proteins to toxoids in an attempt to reduce the frequency of adverse reactions in vaccination programmes, it is preferable to purify toxins before treatment with formaldehyde.