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author:("nfret, alone")
1.  Yellow fever vaccine induces integrated multilineage and polyfunctional immune responses 
The Journal of Experimental Medicine  2008;205(13):3119-3131.
Correlates of immune-mediated protection to most viral and cancer vaccines are still unknown. This impedes the development of novel vaccines to incurable diseases such as HIV and cancer. In this study, we have used functional genomics and polychromatic flow cytometry to define the signature of the immune response to the yellow fever (YF) vaccine 17D (YF17D) in a cohort of 40 volunteers followed for up to 1 yr after vaccination. We show that immunization with YF17D leads to an integrated immune response that includes several effector arms of innate immunity, including complement, the inflammasome, and interferons, as well as adaptive immunity as shown by an early T cell response followed by a brisk and variable B cell response. Development of these responses is preceded, as demonstrated in three independent vaccination trials and in a novel in vitro system of primary immune responses (modular immune in vitro construct [MIMIC] system), by the coordinated up-regulation of transcripts for specific transcription factors, including STAT1, IRF7, and ETS2, which are upstream of the different effector arms of the immune response. These results clearly show that the immune response to a strong vaccine is preceded by coordinated induction of master transcription factors that lead to the development of a broad, polyfunctional, and persistent immune response that integrates all effector cells of the immune system.
doi:10.1084/jem.20082292
PMCID: PMC2605227  PMID: 19047440
2.  Standardization of cytokine flow cytometry assays 
BMC Immunology  2005;6:13.
Background
Cytokine flow cytometry (CFC) or intracellular cytokine staining (ICS) can quantitate antigen-specific T cell responses in settings such as experimental vaccination. Standardization of ICS among laboratories performing vaccine studies would provide a common platform by which to compare the immunogenicity of different vaccine candidates across multiple international organizations conducting clinical trials. As such, a study was carried out among several laboratories involved in HIV clinical trials, to define the inter-lab precision of ICS using various sample types, and using a common protocol for each experiment (see additional files online).
Results
Three sample types (activated, fixed, and frozen whole blood; fresh whole blood; and cryopreserved PBMC) were shipped to various sites, where ICS assays using cytomegalovirus (CMV) pp65 peptide mix or control antigens were performed in parallel in 96-well plates. For one experiment, antigens and antibody cocktails were lyophilised into 96-well plates to simplify and standardize the assay setup. Results (CD4+cytokine+ cells and CD8+cytokine+ cells) were determined by each site. Raw data were also sent to a central site for batch analysis with a dynamic gating template.
Mean inter-laboratory coefficient of variation (C.V.) ranged from 17–44% depending upon the sample type and analysis method. Cryopreserved peripheral blood mononuclear cells (PBMC) yielded lower inter-lab C.V.'s than whole blood. Centralized analysis (using a dynamic gating template) reduced the inter-lab C.V. by 5–20%, depending upon the experiment. The inter-lab C.V. was lowest (18–24%) for samples with a mean of >0.5% IFNγ + T cells, and highest (57–82%) for samples with a mean of <0.1% IFNγ + cells.
Conclusion
ICS assays can be performed by multiple laboratories using a common protocol with good inter-laboratory precision, which improves as the frequency of responding cells increases. Cryopreserved PBMC may yield slightly more consistent results than shipped whole blood. Analysis, particularly gating, is a significant source of variability, and can be reduced by centralized analysis and/or use of a standardized dynamic gating template. Use of pre-aliquoted lyophilized reagents for stimulation and staining can provide further standardization to these assays.
doi:10.1186/1471-2172-6-13
PMCID: PMC1184077  PMID: 15978127
3.  A Canadian national survey of attitudes and knowledge regarding preventive vaccines 
Background
Vaccines have virtually eliminated many diseases, but public concerns about their safety could undermine future public health initiatives.
Objective
To determine Canadians' attitudes and knowledge about vaccines, particularly in view of increasing public concern about bioterrorism and the possible need for emergency immunizations after weaponized anthrax incidents and the events of September 11, 2001.
Method
A 20-question survey based on well-researched dimensions of vaccine responsiveness was telephone-administered to a random sample of N = 1330 adult Canadians in January, 2002.
Results
1057 (79.5%) completed the survey. Respondents perceived vaccines to be highly effective and demonstrated considerable support for further vaccine research. However, results also indicate a lack of knowledge about vaccines and uncertainty regarding the safety.
Conclusions
Support for vaccines is broad but shallow. While Canadians hold generally positive attitudes about vaccines, support could be undermined by widely publicized adverse events. Better public education is required to maintain support for future public health initiatives.
doi:10.1186/1476-8518-1-3
PMCID: PMC280696  PMID: 14613575
preventive vaccines; attitudes; knowledge; nationwide Canadian survey

Results 1-3 (3)