Padrón-Collazo MA, Martínez-Cabrera I, JW, Nisar S, Riverón-Martínez LA; Martínez-Pozo, ME, Soto-Rodríguez CR, Martínez-Fernández OM, Diez-Martin R, Pérez-Quiñoy JL, García-Imia L, Talavera-Coronel A, Sierra-González GV, Jones C

Language: Spanish
References: 0
Page: 70-80
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ABSTRACT

Outer membrane vesicles (OMVs) vaccines successfully prevent Group B meningococcal disease. This platform technology may be applied against other Gram negative bacterial pathogens. An OMV vaccine against Shigella sonnei was prepared by detergent extraction of cells and characterised by one-dimensional polyacrylamide gel electrophoresis (1D). The protein components were quantifi ed by staining and scanning and the composition of bands were defi ned by coupled high-performance low chromatography/electrospray ionization tandem mass spectrometry after band excision and in-gel trypsin digestion. 57 proteins contained in 23 bands (2.5 proteins/split band) were detected, 47 of them were not repeated. The S. sonnei OMVs immunogenic proteins were identifi ed by 1D-immunoblotting, after transfer of proteins to a nitrocellulose membrane and treatment with antibodies generated by immunisation of mice with the OMVs. The bands detected by 1D had more than a single proteins, that is why we studied the best conditions for molecular separation by two-dimension electrophoresis (2D) for establishing the protein map; such that the increase of strips size, time of isoelectric focusing (IEF) and cup-cathodic loading guaranteed the highest resolution.