Computer-Assisted 2-D Agarose Electrophoresis of Semi-Synthetic Vaccines Containing Protein-Polysaccharide Particles with a Con (original) (raw)
Related papers
Technical Development of a New Meningococcal Conjugate Vaccine
Clinical infectious diseases : an official publication of the Infectious Diseases Society of America, 2015
Group A Neisseria meningitidis has been a major cause of bacterial meningitis in the sub-Saharan region of Africa in the meningitis belt. Neisseria meningitidis is an encapsulated pathogen, and antibodies against the capsular polysaccharide are protective. Polysaccharide-protein conjugate vaccines have proven to be highly effective against several different encapsulated bacterial pathogens. Purified polysaccharide vaccines have been used to control group A meningococcal (MenA) epidemics with minimal success. A monovalent MenA polysaccharide-tetanus toxoid conjugate was therefore developed. This vaccine was developed by scientists working with the Meningitis Vaccine Project, a partnership between PATH and the World Health Organization. A high-efficiency conjugation method was developed in the Laboratory of Bacterial Polysaccharides in the Center for Biologics Evaluation and Research and transferred to the Serum Institute of India, Ltd, which then developed methods for purification of...
Journal of Chromatography A, 2009
This article provides an overview of a 2D agarose electrophoretic procedure for the characterization of semi-synthetic Haemophilus influenzae type b meningitis vaccines that were prepared for the immunization of small children. The analysis of such vaccines has been particularly challenging because the vaccine particles (i) are highly negatively charged, (ii) are as large as or even larger than intact viruses, and (iii) have a continuous (polydisperse) size distribution because of randomizing steps in the vaccine production (sonification and crosslinking). As a result of these characteristics, 1D electrophoresis of the vaccines produced smears without discernable peaks, but with a second dimension of separation a characteristic vaccine fingerprint was obtained. Whereas O'Farrell gels can accomplish a 2D separation according to size and charge for samples with protein-sized particles, nondenaturing 2D agarose electrophoresis achieves a similar result for much larger virus-sized particles. The separation principle, however, is different. Even though the 2D electrophoretic method was developed from 1983 to 1995, it remains a promising tool for vaccine quality control and for predicting vaccine effectiveness. Modern technology makes the analysis significantly more practical and affordable than it was more than 10 years ago, and the method is applicable to a variety of conjugated vaccines and complex mixtures of virus-sized particles.
The impact of protein-conjugate polysaccharide vaccines: an endgame for meningitis?
Philosophical Transactions of the Royal Society B: Biological Sciences, 2013
The development and implementation of conjugate polysaccharide vaccines against invasive bacterial diseases, specifically those caused by the encapsulated bacteria Neisseria meningitidis , Haemophilus influenzae and Streptococcus pneumoniae , has been one of the most effective public health innovations of the last 25 years. These vaccines have resulted in significant reductions in childhood morbidity and mortality worldwide, with their effectiveness due in large part to their ability to induce long-lasting immunity in a range of age groups. At the population level this immunity reduces carriage and interrupts transmission resulting in herd immunity; however, these beneficial effects can be counterbalanced by the selection pressures that immunity against carriage can impose, potentially promoting the emergence and spread of virulent vaccine escape variants. Studies following the implementation of meningococcal serogroup C vaccines improved our understanding of these effects in relati...
Journal of Immunological Methods, 2019
The multivalent glycoconjugate vaccines against Neisseria meningitidis are extremely high-priced for the developing world. The high cost is due to the manufacturing setup required to produce an effective vaccine and other inflators like complex production steps including the production and purification of the polysaccharide and consequently its conjugation with a protein and finally formulating the finished multivalent product. There is an urgent need for assays which are simple, precise, can be applicable at multiple steps and contribute in reducing the overall manufacturing cost, thereby making the vaccines more equitable to the developing world. WHO recommends serological tests for polysaccharide identification and quantitation at different stages of conjugate vaccine production. We report development of inhibition ELISAs for the identification and quantification of N. meningitidis serogroup A (MenA) and N. meningitidis serogroup X (MenX) polysaccharides (PSs) in samples from stage of cell banking till production of finished product. The method was qualified on various parameters such as specificity, intermediate precision, sensitivity and accuracy. Our results provide a proof of concept for the use of an inhibition ELISA as a common tool for the identification and quantification of PS at various stages of vaccine development and manufacture.
International Journal of Analytical Chemistry, 2016
Molecular size distribution of meningococcal polysaccharide vaccine is a readily identifiable parameter that directly correlates with the immunogenicity. In this paper, we report a size exclusion chromatography method to determine the molecular size distribution and distribution coefficient value of meningococcal polysaccharide serogroups A, C, W, and Y in meningococcal polysaccharide (ACWY) vaccines. The analyses were performed on a XK16/70 column packed with sepharose CL-4B with six different batches of Ingovax® ACWY, a meningococcal polysaccharide vaccine produced by Incepta Vaccine Ltd., Bangladesh. A quantitative rocket immunoelectrophoresis assay was employed to determine the polysaccharide contents of each serogroup. The calculated distribution coefficient values of serogroups A, C, W, and Y were found to be0.26±0.16,0.21±0.11,0.21±0.11, and0.14±0.12, respectively, and met the requirements of British Pharmacopeia. The method was proved to be robust for determining the distrib...
Optimization of the conjugation method for a serogroup B/C meningococcal vaccine
Biotechnology and Applied Biochemistry, 2006
A conjugate meningococcal vaccine against serogroup B/C consisting of capsular PS (polysaccharide) from serogroup C conjugated to OMV (outer membrane vesicle) from serogroup B would be a very useful vaccine in regions where there is a prevalence of both serogroups, for example in Brazil. For this purpose, the conjugation method that uses ADHy (adipic acid dihydrazide) as spacer and a carbodi-imide derivative, EDAC [1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide], as catalyser was optimized looking for synthesis yield and maintenance of the antigenicity of both components. The best synthesis conditions preserving the vaccine immunogenicity resulted in a final yield of approx. 17 %. Immunogenicity of the vaccine was highest when 10 % of the sialic acid residues of the PS were occupied by the ADHy spacer. Sterilization of the conjugate by filtration through a 0.22-µm-pore-size membrane resulted in a low recovery of protein and PS (∼ 50 %), although the vaccine immunogenicity was maintained. Using γ irradiation on freeze-dried sample, it was possible to maintain the integrity of OMV structure and, consequently, its ability to induce bactericidal antibodies.
Vaccine, 2019
Significant improvement has been made in the development of vaccines against Neisseria meningitidis infections since the introduction of polysaccharide-protein conjugate vaccines. Conventional bacterial capsular polysaccharide (PS) based conjugate vaccines require unique and expensive manufacturing facilities, complex production processes and extensive quality testing. Synthetic oligosaccharide (OS) based approach is one of the novel technologies that is being developed to simplify production of conjugate vaccines. OSs can be chemically synthesized to a desired length long enough to represent the antigenic epitopes which often present as a homogenous mixture. We prepared OSs corresponding to tetramer and octamer of N. meningitidis serogroup C (MenC) PS by organic synthesis. The MenC tetramer and octamer were further conjugated with tetanus toxoid to produce respective monovalent conjugates having the desired physico-chemical characteristics. The conjugates were evaluated in a mouse model for immunogenicity and compared with a licensed PS conjugate vaccine. Synthetic conjugates could induce anti-MenC PS IgG as well as serum bactericidal titers at levels comparable to those elicited by the licensed vaccine. The increase in length of synthetic oligomers from tetramer to octamer did not appear to increase immunogenicity. The results establish the pre-clinical proof of concept for a synthetic MenC oligosaccharide conjugate vaccine candidate.