Quality Control Study Of Residual Materials In Hib Polysaccharide Protein Conjugate Vaccine

Haemophilus influenza type b conjugates vaccine is usually called Hib conjugate vaccine. Once people especially children under2years old are vaccinated by it, the vaccine could produce specific protected antibodies against Haemophilus influenzae type B. The composition of Hib capsular antigen is polyribosylribitol phosphate (PRP), which is poorly immunogenicand belongs to the T cell independent antigen. The immune system of infants and children under2years old is not yet mature, the Hib capsular polysaccharide antigen should be set from T cell independent into T cell dependent antigen, therefore, chemical means to build covalent combination between protein molecules and Hib capsule polysaccharide are needed to make the capsular polysaccharide obtain carrier effect. Finally the vaccine getthe enhanced immune effect after two doses of immunization.This paper consists of three parts. The first part is the preface, which introduces the background information of Hib vaccine and the main production process, then reviews the chemical coupling methods for polysaccharide-protein conjugate vaccine commonly used in the manufacturing process. The second part is the research report, in which four analytical methods are set up for cyanogen bromide (CNBr),1-cyano-4-dimethylamino pyridinium tetrafluoroborate (CDAP), carbodiimide (EDAC) and adipic acid hydrazide (ADH) owing to these four substances commonly used in conjugation chemistry, the mthods set up here could meet the requirements, the details are as below:i. A detection method of residual cyanogen bromide in Hib conjugate vaccine are set up. The method is based on a high performance anion exchange chromatography with electrochemical detector, and a single analysis cycle is only10minutes. The method was validated and the method is specific. The LOD can be achieved 5ug/Lwith the, linear detection range of10-100μg/Lⅱ. A detection method of residual CDAP in Hib conjugate vaccine is set up, which is based on the high performance cation exchange chromatography with UV detector.An analysis cycle of the method need only30minutes. The results show that the method is specific with LOD of16μg/L and the linear detection range of100-1000μg/L.iii. A detection method of residual EDAC in Hib conjugate vaccine is set up, the equipment used in this method is a high performance liquid chromatography and three triple quadrupole mass spectrometer. The method is specific and the LOD is0.2μg/L with the detection linear range of10-100μg/L. Using this method just5-8minutes is needed for a single sample analysis. The results show that the method is simple, accurate and reliable.iv. A colorimetric spectrophotometric detection method is set up for residual ADH in Hib conjugate vaccine The method, is time saving and, simple operation and low cost. The results showed that, the method is specific and linear detection range is2-20μg/mL with LOD of1μg/mL.The third part is the conclusion of the study, prospects and the summary of experiences. Four detection methods established in this study are effective and reliable, and can be used to make quantitative analysis for the residual related substances, providing methodological support validation for the production process, quality control for Hib combined vaccine and further improvement of the vaccine quality. In the field of polysaccharide-protein conjugate vaccine, different combination methods require different chemical substances, therefore, in order to ensure the safety of the vaccine, there are still a lot of work be carried out in the future.