Heterobifunctional reagent, N-succinimidyl 3-(2-pyridylthio) propionate (SPDP) was used for synthesis of polysaccharide and protein conjugates that could be considered for human use. Phosphocholine (PC) specific antibodies elicited by unencapsulated pneumococci confer protection against encapsulated pneumococci. The cell wall polysaccharide (Cw-Ps) is the most likely structure reacting with these PC antibodies. The Cw-Ps was conjugated to bovine serum albumin using SPDP. Rabbits immunized with the conjugate synthesized antibodies toward the Cw-Ps backbone; no PC antibodies were detected. The rabbit antiserum did not confer protection against pneumococci types 3 and 6A in mouse model. Species-specific protection conferred by other PC binding antibodies could not be absorbed with Cw-Ps. Other cell surface PC-containing antigens will be sought using antiserum specific to Cw-Ps to remove this structure. The Vi capsular polysaccharide (ViCPS) is a potential protective antigen of Salmonella typhi, the causative agent of typhoid fever. ViCPS was thiolated with cystamine and then conjugated with a carrier protein through SPDP. Mice immunized with ViCPS conjugate elicited five to ten times higher antibody level than with ViCPS alone; a booster effect was also observed with the conjugate. A clinical study in adult volunteers using cholera toxin or diphtheria toxoid as a carrier protein is planned. The final product using SPDP contains a disulfide bond. Other cross-linking reagents of maleimide or a halogen will be studied for their ability to form a conjugate without a disulfide bond.
