Investigation of recombinant Streptococcus pneumoniae vaccine antigen adsorbed onto an aluminum adjuvant.

摘要

A chemiluminescent nitrogen detector (CLND) has been evaluated for determining the concentration of an aluminum-adsorbed recombinant Streptococcus pneumoniae (Sp) vaccine antigen. Quantification of the antigen was based upon several nitrogen-containing compounds used to calibrate the CLND. All calibrants (6.75--400 mug/mL) generated linear standard curves, with slopes being directly proportional to the % nitrogen. The CLND was capable of analyzing antigen-adjuvant suspensions (adsorbed + unbound antigen) without any sample pre-treatment. To measure unbound antigen, the suspension was centrifuged and an aliquot of supernatant removed for analysis; the difference between these two measurements was the amount of adsorbed antigen. This methodology provides an indirect measurement of the concentration of a vaccine antigen adsorbed onto an aluminum adjuvant.; The adsorption of three Sp vaccine antigens by aluminum-containing adjuvants was then studied using the CLND. The antigens showed high binding affinity with aluminum hydroxide adjuvant described by the Langmuir equation but virtually no binding to aluminum phosphate adjuvant. A number of conditions were evaluated to determine whether electrostatic or hydrophobic interactions were responsible for the observed binding, but no significant change in the adsorptive capacity was observed. This indicates that neither electrostatic nor hydrophobic interactions appear to be responsible for the observed binding, which means that ligand exchange may be the primary mechanism for this interaction. Further binding competition studies resulted in evidence that Sp antigen follows the trend that proteins with higher adsorptive coefficients are able to displace those with lower adsorptive coefficients.; Finally, the impact on immunogenicity of the degree of adsorption of three Sp vaccine antigens to aluminum adjuvants was studied. The three antigens evaluated were highly adsorbed by aluminum hydroxide adjuvant, but not adsorbed by aluminum phosphate adjuvant. All of the Sp antigens adjuvanted with aluminum hydroxide gave higher antibody responses than formulations prepared with aluminum phosphate or non-adjuvanted antigen. Varying the percent aluminum-bound Sp antigen in the formulated vaccine affected the observed antibody responses. It was concluded from these observations that the antibody response observed for Sp antigens in this study is stimulated by a depot effect of the antigen bound to an aluminum adjuvant.