Activity in saline of phthalylated or succinylated derivatives of mycobacterial water-soluble adjuvant.
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A water-soluble fraction (WSA) of the cell wall can substitute for mycobacterial cells in Freund complete adjuvant. However, when WSA is administered in saline instead of in a water-in-oil emulsion, its adjuvant activity is very weak, and under certain experimental conditions it can even inhibit the humoral immune response. The data reported in the present study show that after treatment by phthalic or succinic anhydride the adjuvant activity of WSA was markedly changed, since high levels of circulating antibodies were produced when these derivatives were administered with an antigen in an aqueous medium. Moreover, the antigenic determinants of WSA were modified and acylated WSA had no tuberculin-like activity. (+info)
Immunoglobulin-specific radioimmunoprecipitation assays for quantitation of nasal secretory antibodies to hemagglutinin of type A influenza viruses.
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Radioimmunoprecipitation (RIP) assays were developed to selectively quantitate class-specific antibodies to purified hemagglutinins (HA) of type A influenza virus in nasal secretions. Rabbit anti-human secretory piece of immunoglobulin A (IgA) and rabbit anti-human IgG were used as second antibodies. A third antibody, goat anti-rabbit IgG, was incorporated into the system to separate immune complexes formed between iodinated HA, nasal wash test specimen, and second antibody. The utilization of this reagent avoided the need for large quantities of IgA and IgG antibody-negative carrier secretions. Nasal was specimens obtained from 14 adults immunized with an inactivated type A influenza virus vaccine were evaluated by RIP and viral neutralization assays. Significant homologous postvaccination secretory IgA and IgG antibody levels were demonstrable in 13 (93%) of individuals by RIP, whereas only 5 (36%) exhibited rises by viral neutralization tests. Moreover, the geometric mean IgA and IgG antibody levels were at least 20- and 37-fold greater than the neutralizing antibody titer. The pattern of heterologous immunoglobulin-specific antibody responses tended to be similar to those observed with the homologous HA subunit. (+info)
Potential advantages of DNA immunization for influenza epidemic and pandemic planning.
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Immunization with purified DNA is a powerful technique for inducing immune responses. The concept of DNA immunization involves insertion of the gene encoding the antigen of choice into a bacterial plasmid and injection of the plasmid into the host where the antigen is expressed and where it induces humoral and cellular immunity. The most effective routes and methods for DNA immunization are bombardment with particles coated with DNA ("gene gun" technique), followed by the intramuscular and intradermal routes. DNA immunization technology has the potential to induce immunity to all antigens that can be completely encoded in DNA, which therefore include all protein, but not carbohydrate, antigens. DNA immunization results in presentation of antigens to the host's immune system in a natural form, like that achieved with live-attenuated vaccines. The DNA immunization strategy has the potential to rapidly provide a new vaccine in the face of an emerging influenza pandemic. (+info)
Mucosal immunity to influenza without IgA: an IgA knockout mouse model.
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IgA knockout mice (IgA-/-) were generated by gene targeting and were used to determine the role of IgA in protection against mucosal infection by influenza and the value of immunization for preferential induction of secretory IgA. Aerosol challenge of naive IgA-/- mice and their wild-type IgA+/+ littermates with sublethal and lethal doses of influenza virus resulted in similar levels of pulmonary virus infection and mortality. Intranasal and i.p. immunization with influenza vaccine plus cholera toxin/cholera toxin B induced significant mucosal and serum influenza hemagglutinin-specific IgA Abs in IgA+/+ (but not IgA-/-) mice as well as IgG and IgM Abs in both IgA-/- and IgA+/+ mice; both exhibited similar levels of pulmonary and nasal virus replication and mortality following a lethal influenza virus challenge. Monoclonal anti-hemagglutinin IgG1, IgG2a, IgM, and polymeric IgA Abs were equally effective in preventing influenza virus infection in IgA-/- mice. These results indicate that IgA is not required for prevention of influenza virus infection and disease. Indeed, while mucosal immunization for selective induction of IgA against influenza may constitute a useful approach for control of influenza and other respiratory viral infections, strategies that stimulate other Igs in addition may be more desirable. (+info)
Influenza vaccination among the elderly in Italy.
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This article surveys the attitudes and perceptions of a random sample of the elderly population in three regions of Italy on the use and efficacy of influenza vaccine. The data were collected by direct interviews using a standard questionnaire. The results show that vaccination coverage against influenza is inadequate (26-48.6%). The major reasons for nonvaccination were lack of faith in the vaccine and disbelief that influenza is a dangerous illness. These data emphasize the need for a systematic education programme targeted at the elderly and the provision of influenza vaccination, with the increased cooperation of general practitioners. (+info)
Protection of mice against a lethal influenza virus challenge after immunization with yeast-derived secreted influenza virus hemagglutinin.
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The A/Victoria/3/75 (H3N2-subtype) hemagglutinin (HA) gene was engineered for expression in Pichia pastoris as a soluble secreted molecule. The HA cDNA lacking the C-terminal transmembrane anchor-coding sequence was fused to the Saccharomyces cerevisiae alpha-mating factor secretion signal and placed under control of the methanol-inducible P. pastoris alcohol oxidase 1 (AOX1) promoter. Growth of transformants on methanol-containing medium resulted in the secretion of recombinant non-cleaved soluble hemagglutinin (HA0s). Remarkably, the pH of the induction medium had an important effect on the expression level, the highest level being obtained at pH 8.0. The gel filtration profile and the reactivity against a panel of different HA-conformation specific monoclonal antibodies indicated that HA0s was monomeric. Analysis of the N-linked glycans revealed a typical P. pastoris type of glycosylation, consisting of glycans with 10-12 glycosyl residues. Mice immunized with purified soluble hemagglutinin (HA0s) showed complete protection against a challenge with 10 LD50 of mouse-adapted homologous virus (X47), whereas all control mice succumbed. Heterologous challenge with X31 virus [A/Aichi/2/68 (H3N2-subtype)], resulted in significantly higher survival rates in the immunized group compared with the control group. These results, together with the safety, reliability and economic potential of P. pastoris, as well as the flexibility and fast adaptation of the expression system may allow development of an effective recombinant influenza vaccine. (+info)
Measuring the effects of reminders for outpatient influenza immunizations at the point of clinical opportunity.
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OBJECTIVE: To evaluate the influence of computer-based reminders about influenza vaccination on the behavior of individual clinicians at each clinical opportunity. DESIGN: The authors conducted a prospective study of clinicians' influenza vaccination behavior over four years. Approximately one half of the clinicians in an internal medicine clinic used a computer-based patient record system (CPR users) that generated computer-based reminders. The other clinicians used traditional paper records (PR users). MEASUREMENTS: Each nonacute visit by a patient eligible for an influenza vaccination was considered an opportunity for intervention. Patients who had contraindications for vaccination were excluded. Compliance with the guideline was defined as documentation that a clinician ordered the vaccine, counseled the patient about the vaccine, offered the vaccine to a patient who declined it, or verified that the patient had received the vaccine elsewhere. The authors calculated the proportion of opportunities on which each clinician documented action in the CPR and PR user groups. RESULTS: The CPR and PR user groups had different baseline compliance rates (40.1 and 27.9 per cent, respectively; P<0.05). Both rates remained stable during a two-year baseline period (P = 0.34 and P = 0.47, respectively). The compliance rates in the CPR user group increased 78 per cent from baseline (P<0.001), whereas the rates for the PR user group did not change significantly (P = 0.18). CONCLUSIONS: Clinicians who used a CPR with reminders had higher rates of documentation of compliance with influenza-vaccination guidelines than did those who used a paper record. Measurements of individual clinician behavior at the point of each clinical opportunity can provide precise evaluation of interventions that are designed to improve compliance with guidelines. (+info)
Detection of intracellular antigen-specific cytokines in human T cell populations.
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Determination of antigen-specific cytokine responses of T lymphocytes after vaccination is made difficult by the low frequency of responder cells. In order to detect these responses, the profile of intracellular cytokines was analyzed using flow cytometry after antigenic expansion. Peripheral blood mononuclear cells were stimulated with antigens for 5 days, further expanded with interleukin (IL)-2, and then restimulated on day 10. Cytokine production was detected by intracellular staining with monoclonal antibodies after saponin-based permeabilization. Influenza expansion resulted in specific interferon-gamma (IFN-gamma) production of 6%-20%, with less IL-4 production (0%-2%). Tetanus toxoid resulted in even greater production. IL-4 and IFN-gamma were produced mainly by memory cells of the CD45RO+ phenotype. IFN-gamma production was contributed by both CD4 and CD8 populations. These methods were then applied to a clinical trial of a candidate human immunodeficiency virus type 1 vaccine. Antigen-specific increases in IFN-gamma were measured, which corresponded to antibody production, lymphoproliferation, and skin testing. (+info)