Dot blot para identidad del polisacárido de Streptococcus pneumoniae serotipo 14 y toxoide tetánico en vacunas conjugadas

Regulatory authorities recommend the use of immunochemical techniques to determine the identity of the antigens present on the conjugate vaccines. With the emergence of multivalent vaccines, it has become necessary to use immunochemical techniques using monoclonal antibodies to increase the sensitivity in the determination. The aim of this study was to establish the analytical conditions that would allow to use the Dot Blot as a technique to determine the identity of capsular polysaccharides from Streptococcus pneumoniae serotype 14 (PsC 14) and the tetanus toxoid (TT) used as carrier protein in multivalent conjugate vaccines. The MAb against the PsC 14 and TT were used; besides were used some batchs of monovalent conjugates of PsC 14; batchs of a heptavalent conjugate pneumococcal vaccine candidate with TT as carrier protein and batchs of TT, all produced at Finlay Institute of Vaccines. The results showed that for the determination of the antigenic identity were optimal volumes of 10µL of monovalent conjugate samples at a dilution of 1/10 (vol/vol) and equal volume for heptavalent vaccines. For MAb was demonstrated that 2µg/mL concentration of MAb against the PSC 14 and 2.5µg/ mL for MAb against TT were sufficient to successfully perform the determinations; while incubation times were adjusted at 37oC for 30min. The work allows us to conclude that working conditions in the laboratory were established to determine the identity of the capsular polysaccharides of S. pneumoniae serotype 14 and for tetanus toxoid as carrier protein in the heptavalent conjugate vaccines by Dot Blot method.