ArticleMethods and protocols2026
Single Radial Immunodiffusion (SRID) Assay for Quantitative Determination of Recombinant SOD Protein in a Brucellosis Vaccine Candidate: Method Development and Validation.
Article in Methods and protocols, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
Recombinant superoxide dismutase (SOD) protein is a promising antigen candidate for brucellosis vaccines, and its quantitative determination is essential for vaccine quality control, standardization, and regulatory compliance. Although enzyme-linked immunosorbent assay (ELISA) is widely used for antigen quantification, its application to vaccine formulations may be limited due to chemical modifications of antigenic determinants occurring during detoxification and adjuvantation processes. Therefore, alternative analytical methods, such as single radial immunodiffusion (SRID), which enable direct antigen quantification independent of antigenic determinant modifications, represent valuable tools for vaccine analysis. The aim of this study was to develop and validate a single radial immunodiffusion (SRID) assay for the quantitative determination of recombinant SOD protein and to obtain hyperimmune sera in sheep. Sheep were immunized with recombinant SOD protein formulated with aluminum hydroxide or AddaS03 adjuvants. Antibody responses were evaluated using agar gel immunodiffusion (AGID) and enzyme-linked immunosorbent assay (ELISA). Immunization with aluminum hydroxide induced higher antibody titers than AddaS03. By day 42, ELISA titers reached 1:25,600 in the aluminum hydroxide group compared with 1:3200 in the AddaS03 group. The obtained hyperimmune sera were used for SRID assay development. The optimized SRID assay demonstrated high reproducibility and specificity, with no cross-reactivity against unrelated proteins. A linear relationship was observed between the square of the precipitation ring diameter (D
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