Potentia carrier for conjugate vaccines: OMP-38 protein from Shigella flexneri 3a - isolation and purification strategies

Abstract lntroduction The Enterobacteriaceae family, Gram-negative, rod-shaped bacteria are the maincause of diarrheal diseases.A nnually about 1.9m illion people die of diarrheal diseases,m ainiy in developingcountries, especially among infants and children under 5 years old. The development of vaccines againstenteric pathogens represents a serious challenge, because of raising bacterial antibiotic-resistance and noaccess to diagnostic tests providing pathogen identification and treatment. The OMP-38 may be a potentialcarrier or antigen for conjugate vaccine as an immunogenic and protective component mainly exposed onbacterial cellwall. Aim The aim of this work was to optimize the purification conditions of OMP-38 proteinfrom Shigella flexneri 3a crude extract, to obtain a homogenous protein for vaccine design. Materialsand Methods The strains were obtained from the Polish Collection of Microorganisms of the Institute ofImmunology and Experimental Therapy, Polish Academy of Sciences. Bacteria were grown in BHI broth at37'C with aeration. The outer membrane proteins were extracted according to the method of Arcidiaconoet al. with own modifications. Purification of the OMP-38 was based on chromatographic methods: sizeexclusion chromatography and ion-exchange chromatography using non-ionic detergent Triton X-100. Thefractions containing the OMP-38 were identified by SDS-PAGE according to Laemmli. An immunoblottingand ELISA with human umbilical cord serum were performed to confirm antigenicity of OMP-38. ResultsA convenient and reliable method for the purification of OMP-38 and an assessment of the purity, yield,antigenicity and immunogenicity of the purified protein were developed. Using two step purificationmethod we have obtained a pure, antigenic and immunogenic protein with a satisfactory yield of 1.70 mgoMP-38 per gram of Ęphilized bacterial mass. ConclusionThe purified oMP-38 may cause protectiveimmune response and can be used as a potential antigen or carrier for vaccine design.