Preparation purification and characterization of neisseria meningitidis group a capsular polysaccharide tetanus toxoid conjugates

Conjugation of Neisseria meningitidis A Capsular polysaccharide [CPS] to tetanus toxiod [TT] was aimed at in this work, with the intention of developing a new vaccine which may be more protective, specially to the age group below 18 months. The CPS was prepared and applied to sepharose CL-48 column. Fractions were collected and monitored for their carbohydrate content. They were divided according to their Kd into two pools [low and high molecular weight pools]. Tetanus toxoid was applied to sephacryl S-300 column and fractions were pooled into three pools. The second pool, having a molecular weight of 150 KD, as determined by SDS-PAG was used as the protein carrier. Conjugation was carried out, using cyanogen bromide activated, adipic acid dihydrazide derivative of the CPS. Binding to TT was achieved by a carbodiimide-mediated reaction. The obtained high and low molecular weight conjugates were subjected to gel filtration on sepherose CL-4B column, and the carbohydrate and protein content in the fractions were monitored. In the low molecular weight conjugates, there was a shift in the elution pattern the left. This indicates an increase in the molecular size. Fractions were pooled and the pools were tested for their antigenicity, against a standard horse anticapsular polysaccharide antibodies. The ratio of carbohydrate to protein in the conjugate was also determined

Antibody response against neisseria meningitidis group A capsular polysaccharide in mice receving meningococcal a capsular polysaccharide-tetanus toxoid conjugates

C 57 BL/6J Mice were used to study the antibody response against neisseria meningitidis group A capsular polysaccharide [CPS]. Four groups [8 mice each] were injected twice subcutaneously [SC] with a 2 microgram equivalent of meningococcal A capsular polysaccharide, and a fifth group received an equal volume of saline diluent and served as the control group. ELISA was done using tyraminated CPS preparation to coat the polyvinyl chloride flexible plates. Results showed that the postimmunization sera of the groups receiving either of the previously prepared high and low molecular weight conjugates, or capsular polysaccharide added to tetanus toxoid had, a better antibody titer than their preimmunization serum. The control group had no change in antibody response, while the group receiving the standard CPS vaccine showed vary little increase. However, only the group receiving the low molecular weight conjugate showed a significant increase in titer as compared to the group receiving CPS added to tetanus toxoid. The high molecular weight conjugate group did not show this difference. Furthermore, the sera of low molecular weight conjugate group are partiaty inhibited when preincubated with CPS before the ELISA assay. The results showed that the low molecular weight conjugate is a better vaccine against meningococcus A than either the CPS alone, CPS added to tetanus toxoid or the high molecular weight conjugate