RESUMO
The Dtxd (Diphtheria toxoid) was the first antigen encapsulated within liposomes, their adjuvant properties were discovered (their capacity to enhance the vaccine immunogenicity). The point here is not to propose a new method to prepare this lipossomal vaccine. The central idea is to give new dresses for old vaccines by using classical and well-established liposome preparation method changing only the encapsulation pH and the immunization protocol. The most appropriate method of Dtxd encapsulation within liposome was based on lipid film hydration in 100 mM citrate buffer, pH 4.0. This was accompanied by changes on protein hydrophobicity, observed by CD and fluorescence spectroscopies. Whenever the Dtxd exposed its hydrophobic residues at pH 4.0, it interacted better with the lipossomal (observed by electrophoretic mobility) film than when its hydrophobic residues were buried (pH 9.0). The Dtxd partition coefficient in Triton-X114 and the acrylamide fluorescence quenching were also pH dependent. Both were bigger at pH 4.0 than at pH 9.0. The relationship protein structure and lipid interaction was pH dependent and now it can be easily maximized to enhance encapsulation of antigens in vaccine development. Mice were primed with formulations containing 5 mug of Dtxd within liposomes prepared in pH 4.0 or 7.0 or 9.0. The boosters were done 38 or 138 days after the first immunization. The IgM produced by immediate response of all lipossomal formulations were higher than the control (free protein). The response patterns and the immune maturity were measured by IgG1 and IgG2a titrations. The IgG1 titers produced by both formulations at pH 4.0 and 7.0 were at least 22 higher than those produced by mice injected lipossomal formulation at pH 9.0. When the boosters were done, 138 days after priming the mice produced a IgG2a titer of 29 and the group that received the booster 30 days after priming produced a titer of 25. The strongest antibody production was the neutralizing antibody (245 higher than the control) produced by those mice injected with lipossomal formulation at pH 4.0 with the booster done 138 days after priming. The simple change on lipossomal pH formulation and timing of the booster enhanced both antibody production and selectivity.
Assuntos
Lipossomos , Vacinas/administração & dosagem , Animais , Formação de Anticorpos , Cromatografia Líquida de Alta Pressão , Dicroísmo Circular , Feminino , Fluorescência , Concentração de Íons de Hidrogênio , Camundongos , Vacinas/química , Vacinas/imunologiaRESUMO
The study described in this article was carried out for the purpose of evaluating the protective effects of two stabilizing solutions- sorbitol-gelatin and glutamic acid-lactose- on freeze-dried measles virus (Schwartz strain) with a view to the production of reference preparations in working lots. The effect of storage at -20 C and -70 C on the potency of stabilized virus suspensions, whether or not freeze-dried, was evaluated over a period of 21 months; the samples were concurrently titered for potency with a standard virus obtained from a laboratory authorized by the World Health Organization. After comparing and commenting on the different phases of the study, the types of cells and stabilizing media used, how the suspensions were stored, the titering of the stabilized viruses, etc., the authors conclude that they have demostrated a more satisfactory stabilization (r= -0.01) of freeze-dried virus suspensions stored at -20 C by sorbitol-gelatin, which is therefore regarded as an effective stabilizer for the preparation of freeze-dried reference measles virus in working lots
Assuntos
Excipientes , Vírus do Sarampo , Padrões de Referência , Vacinas ViraisRESUMO
The study described in this article was carried out for the purpose of evaluating the protective effects of two stabilizing solutions- sorbitol-gelatin and glutamic acid-lactose- on freeze-dried measles virus (Schwartz strain) with a view to the production of reference preparations in working lots. The effect of storage at -20 C and -70 C on the potency of stabilized virus suspensions, whether or not freeze-dried, was evaluated over a period of 21 months; the samples were concurrently titered for potency with a standard virus obtained from a laboratory authorized by the World Health Organization. After comparing and commenting on the different phases of the study, the types of cells and stabilizing media used, how the suspensions were stored, the titering of the stabilized viruses, etc., the authors conclude that they have demostrated a more satisfactory stabilization (r= -0.01) of freeze-dried virus suspensions stored at -20 C by sorbitol-gelatin, which is therefore regarded as an effective stabilizer for the preparation of freeze-dried reference measles virus in working lots
