The content of Haemophilus b polysaccharide (PS) in Haemophilus b-conjugate vaccines in final containers is usally determinened by measuring its ribose using colorimetric orcinol assay. Hoewever, the MSD vaccine contains a large quantity of lactose, a stabilizer, which makes the assay impossible. Rate nephelometry, which is based on the rate of antigen-antibody aggregation, is currently used for the indirect measurement of the Haemophilus b PS content of the vaccines. A direct physico-chemical method to quantitate the PS content has been explored. The results obtained from ongoing experiments showed that the amounts of Haemophilus b PS in the MSD vaccine as well as Connaught and Praxis vaccines could be measured on a Dionex carbohydrate analyzer as follows. The PS in the conjugate vaccine was first depolymerized in strong alkali into its single repeating unit (ribose-ribitol-phosphate) which accounted for 80% of PS if an Haemophilus b PS was used. The PS repeating unit in the alkali-treated sample was then separated from the huge lactose peak by an HPLC column and quantitated. Experiments are planned to measure the amounts of the conjugated and unconjugated PS in the vaccines.
