Recombinant Opc meningococcal protein, folded in vitro, elicits bactericidal antibodies after immunization

Musacchio, Andrea; Carmenate, Tania; Delgado, Maité; González, Sonia

doi:10.1016/s0264-410x(96)00198-3

Cited by 19 publications

(15 citation statements)

References 20 publications

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“…These results show that phenol extraction alone could not efficiently purify aRNA probably due to the high concentration of proteins such as spermidine in the reaction buffer. The addition of guanidinium salt with its chaotropic and denaturing effects [10], improved the purification without compromising yield.…”

Section: Resultsmentioning

confidence: 99%

Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

et al. 2004

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Background: Expression microarrays have evolved into a powerful tool with great potential for clinical application and therefore reliability of data is essential. RNA amplification is used when the amount of starting material is scarce, as is frequently the case with clinical samples. Purification steps are critical in RNA amplification and labelling protocols, and there is a lack of sufficient data to validate and optimise the process.

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Section: Resultsmentioning

confidence: 99%

Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

et al. 2004

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“…Following solubilization in the presence of urea and immunization using Freund's adjuvant, which is not suitable for human use, a bactericidal effect against the Opc ϩϩ homologous strain was reported. Reduced activity was seen with additional Opc ϩ strains, although the sequence of the protein in these heterologous strains was not reported (19).…”

Section: Fig 2 Variation In Antibody Recognition Ofmentioning

confidence: 95%

“…The more conserved nature of Opc, combined with its important role in pathogenesis (29,30) and as a target for bactericidal antibodies (23), suggests its potential for studies as a recombinant vaccine. In a previous study, Musacchio and colleagues (19) reported the expression of recombinant Opc protein in E. coli as a fusion protein with a fragment of another meningococcal protein. Following solubilization in the presence of urea and immunization using Freund's adjuvant, which is not suitable for human use, a bactericidal effect against the Opc ϩϩ homologous strain was reported.…”

Section: Fig 2 Variation In Antibody Recognition Ofmentioning

confidence: 99%

“…A previous study concluded that Opc protein "appears to deserve more attention as a vaccine candidate" (19). While it should be recognized that experiments with laboratory animals may not reproduce precisely the human immune response, the lower immunogenicity of the protein, combined with the effects of variation in both sequence and levels of expression, suggests that it is unlikely to provide effective immunity alone.…”

Section: Fig 2 Variation In Antibody Recognition Ofmentioning

confidence: 99%

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Immunization with Recombinant Opc Outer Membrane Protein from Neisseria meningitidis : Influence of Sequence Variation and Levels of Expression on the Bactericidal Immune Response against Meningococci

et al. 2001

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The opc gene from Neisseria meningitidis was cloned into the pRSETA vector, and recombinant protein was expressed at high levels in Escherichia coli. The protein was readily purified by affinity chromatography and used for immunization with conventional Al(OH) 3 adjuvant or after incorporation into liposomes and Zwittergent micelles. The resulting sera were analyzed for their ability to recognize purified recombinant protein and "native" protein in an enzyme immunoassay with outer membranes and by whole-cell immunofluorescence. Immunization with Al(OH) 3 induced high levels of antibodies which reacted with the purified protein but did not recognize whole cells. In contrast, liposomes and micelles induced antibodies which reacted with the native protein in whole cells. The addition of monophosphoryl lipid A (MPLA) to either liposomes or micelle preparations increased the magnitude of the immune response and induced a wider range of immunoglobulin subclasses. This was associated with the ability of the sera to induce complement-mediated killing of the homologous strain. The most effective bactericidal activity was observed with Opc protein incorporated into liposomes containing MPLA. The magnitude of the bactericidal effect was strongly influenced by the level of expression of the Opc protein and was abolished by limited variation in the sequence of the protein expressed by heterologous strains.

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“…However, OpaJ does not induce bactericidal antibodies in mice [37]. Moreover, bactericidal OpcA antibodies are not effective with target strains expressing negligible OpcA levels [38][39][40], as applied in our bactericidal assay. Based on the lack of bactericidal activity of the opacity proteins and the similar Omp85 levels in the two wt OMVs, no distinction will be made below between the wt 1 and wt 2 control OMVs.…”

Section: Recombinant and Control Omv Vaccinesmentioning

confidence: 95%

Meningococcal Omp85 in Detergent‐Extracted Outer Membrane Vesicle Vaccines Induces High Levels of Non‐Functional Antibodies in Mice

et al. 2013

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The vaccine potential of meningococcal Omp85 was studied by comparing the immune responses of genetically modified deoxycholate-extracted outer membrane vesicles, expressing five-fold higher levels of Omp85, with wild-type vesicles. Groups (n = 6-12) of inbred and outbred mouse strains (Balb/c, C57BL/ 6, OFI and NMRI) were immunized with the two vaccines, and the induced antibody levels and bactericidal and opsonic activities measured. Except for Balb/ c mice, which were low responders, the genetically modified vaccine raised high Omp85 antibody levels in all mouse strains. In comparison, the wild-type vaccine gave lower antibody levels, but NMRI mice responded to this vaccine with the same high levels as the modified vaccine in the other strains. Although the vaccines induced strain-dependent Omp85 antibody responses, the mouse strains showed high and similar serum bactericidal titres. Titres were negligible with heterologous or PorA-negative meningococcal target strains, demonstrating the presence of the dominant bactericidal PorA antibodies. The two vaccines induced the same opsonic titres. Thus, the genetically modified vaccine with high Omp85 antibody levels and the wild-type vaccine induced the same levels of functional activities related to protection against meningococcal disease, suggesting that meningococcal Omp85 is a less attractive vaccine antigen.

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Recombinant Opc meningococcal protein, folded in vitro, elicits bactericidal antibodies after immunization

Cited by 19 publications

References 20 publications

Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

Immunization with Recombinant Opc Outer Membrane Protein from Neisseria meningitidis : Influence of Sequence Variation and Levels of Expression on the Bactericidal Immune Response against Meningococci

Meningococcal Omp85 in Detergent‐Extracted Outer Membrane Vesicle Vaccines Induces High Levels of Non‐Functional Antibodies in Mice

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