The HID protein, derived from non-typeable H. influenzae, is a cell surface protein with a molecular weight of approximately 40 kDa. Recombinant strains of Escherichia coli are used to produce HID, which has been employed as a carrier in multivalent pneumococcal conjugate vaccines for most serotypes. Animal studies have demonstrated the immunogenicity of HID. Hyperimmunization of rats with the non-acylated form of HID results in the production of immunoglobulin A (IgA) and immunoglobulin G (IgG) antibodies, which exhibit strong bactericidal activity against various strains of non-typeable H. influenzae.
HID has the characteristics of surface localization, high conservation, wide distribution, and pathogenicity. In addition, HID has a good prospect of preclinical application. Therefore, HID is expected to function as a carrier protein with the antigenic potential to exert dual protective effects against S. pneumoniae and non-typeable H. influenzae.
Fig.1 The "neglected valency" effect – highlighted portion. (Bröker, et al., 2017)
With HID as a carrier protein, CD BioGlyco provides Pneumococcal Polysaccharide conjugation service. Our services are mainly as follows:
The pneumococcal polysaccharide and HIB carrier protein are sonicated and the molecular weight of each molecule is assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), then sterile filter and refrigerate until it begins to perform conjugation experiment. The conjugation experiment is carried out by the principle of chemistry-based polysaccharide conjugation. After the experiment, dialysis and sterile filtration are used for characterization.
The conjugates are characterized by size exclusion chromatography-multi-angle laser light scattering (SEC-MALLS) and SDS-PAGE to verify the successful coupling. Multiple single polysaccharide conjugates are conjugated to obtain the final multivalent vaccine. Vaccines are diluted and refrigerated in sterile vials until used in animal studies.
Establish animal models and conduct group experiments. The HID-pneumococcal polysaccharide conjugate is injected into animals. The number of injections is determined according to factors such as the concentration of the conjugate, injection dose, and purity. Middle ear fluid and nasopharyngeal swabs are then collected. Bacteriological cultures are performed in the laboratory within 24 hours. S. pneumoniae and H. influenzae isolates are further tested for confirmation and serotyping.
Samples for analysis of S. pneumoniae are plated on blood agar. For nasopharyngeal swabs, the assay is performed on selective blood agar containing gentamicin. Recovered pneumococci are typed by suppressing the response using antiserum.
Serum samples are stored at -20°C until masking analysis. Serum anti-pneumococcal IgG concentrations of each serotype in the vaccine are measured by ELISA, using polysaccharides from a certain serotype for a pre-adsorption step to increase the specificity of the assay.
IgG antibodies to HID are measured by classical ELISA using non-lipidated protein D as coating material.
Fig.2 Process of HID-pneumococcal conjugate development. (CD BioGlyco)
At CD BioGlyco, we will work with our customers to define specific requirements and expectations to meet the partnership objectives. We provide Carbohydrate-based Vaccine Development Services using the Glyco™ Vaccine Development Platform. If you are interested in our services, please contact us for more details without any hesitation.
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