Electron beam fragmentation of bacterial polysaccharides as a method of producing oligosaccharides for the preparation of conjugate vaccines

Abstract

End-group mediated conjugation of bacterial polysaccharides (PSs) to carrier proteins containing T-helper cell epitopes renders such polysaccharides immunogenic also in young infants. Optimal construction of such conjugate vaccines requires fragmentation of the PS prior to the coupling reaction. In the present study a general simple and inexpensive method for the fragmentation of PSs is presented. It is based on the irradiation of isolated PSs in an electron beam accelerator. Exposure of isolated pneumococcal capsular polysaccharides (PnPSs) to ionizing radiation resulted in their partial depolymerization in a radiation dose-dependent manner. Radiation, unlike sonication, generated PnPS fragments of molecular size lower than 50 kDa and as small as 1.5 kDa when high radiation doses were used. These PnPS fragments have terminal reducing groups that can be easily used for chemical activation and subsequent coupling to any chosen carrier protein. The radiation-produced PnPS fragments retained their antigenic epitopes, when compared to native, full-size PnPSs as determined by enzyme-linked immunoassay.

Pathogen specific carbohydrate antigen microarrays: a chip for detection of Salmonella O-antigen specific antibodies

“…We do not know the disease history of these healthy controls and we cannot rule out that the control that exhibited somewhat elevated anti-BO titers might have experienced an earlier clinical or subclinical Salmonella serogroup BO infection. The signal difference between the larger oligosaccharides of serogroup BO (9) and DO (15), respectively, was not as prominent as for the corresponding disaccharide antigens. However, signals were detected a lower concentrations for the larger oligosaccharide antigens and this was even more prominent with the complete PS antigen (data not shown).…”

“…Next, we evaluated a S. Typhimurium BO-specific monoclonal antibody (Mab) preparation. This Mab reacted strongly with the larger Salmonella O4 and O12 containing antigens (9,12,15). Weak cross reactivity towards structure 7 was also observed (Fig.…”

“…Binding to the polysaccharide of group BO was also seen. The group BO-specific antiserum bound strongly to the abequosyl-containing compounds (9,12,13,15) related to the immuno-dominant part of the O4 epitope. Weak reactivity with compounds (7) related to the group DO antigens was also seen, most likely caused by a remaining O-antigen 12 reactivity of this antiserum.…”

“…Hence, either total organic synthesis or specific degradations of the native polysaccharides to give chemically defined poly-or oligosaccharide fragments with high specific antigenic activity is feasible [9,10]. The availability of defined carbohydrate antigens has also spurred the development of new, highly sensitive and miniaturized techniques for detecting antigen-antibody reactions.…”

Protection against Streptococcus suis Serotype 2 Infection Using a Capsular Polysaccharide Glycoconjugate Vaccine

“…Ultrasonic irradiation (sonication) was chosen over fragmentation by chemical (60)(61)(62) or enzymatic (63-66) methods to avoid chemical alterations (67). In addition, it is easy to use, is reliable for labile epitopes, and does not require elimination of excess reagents.…”