52 research outputs found

    Removing N-Terminal Sequences in Pre-S1 Domain Enhanced Antibody and B-Cell Responses by an HBV Large Surface Antigen DNA Vaccine

    No full text
    <div><p>Although the use of recombinant hepatitis B virus surface (HBsAg) protein vaccine has successfully reduced global hepatitis B infection, there are still a number of vaccine recipients who do not develop detectable antibody responses. Various novel vaccination approaches, including DNA vaccines, have been used to further improve the coverage of vaccine protection. Our previous studies demonstrated that HBsAg-based DNA vaccines could induce both humoral and CMI responses in experimental animal models. However, one form of the the HBsAg antigen, the large S antigen (HBs-L), expressed by DNA vaccine, was not sufficiently immunogenic in eliciting antibody responses. In the current study, we produced a modified large S antigen DNA vaccine, HBs-L(T), which has a truncated N-terminal sequence in the pre-S1 region. Compared to the original HBs-L DNA vaccine, the HBs-L(T) DNA vaccine improved secretion in cultured mammalian cells and generated significantly enhanced HBsAg-specific antibody and B cell responses. Furthermore, this improved HBsL DNA vaccine, along with other HBsAg-expressing DNA vaccines, was able to maintain predominantly Th1 type antibody responses while recombinant HBsAg protein vaccines produced in either yeast or CHO cells elicited mostly Th2 type antibody responses. Our data indicate that HBsAg DNA vaccines with improved immunogenicity offer a useful alternative choice to recombinant protein-based HBV vaccines, particularly for therapeutic purposes against chronic hepatitis infection where immune tolerance led to poor antibody responses to S antigens.</p> </div

    HBs-specific antibody responses in Balb/C mice receiving DNA vaccine expressing HBs-L or HBs-L(T), or the empty vector as negative control.

    No full text
    <p>A: Temporal HBs-specific antibody responses in immune mouse sera (1∶500 dilution) detected against commercial HBsAg antigen. The arrows show the time points of DNA immunization. The OD values are expressed as the average of group. B and C: HBs-specific IgG titers (reciprocal serum dilution) induced by HBs-L or HBs-L(T) DNA vaccine or empty vector were detected against commercial HBsAg antigen (B) by ELISA or by a commercial HBs-specific antibody detection kit. Antibody titers were measured using sera collected at 2 weeks after the 3<sup>rd</sup> DNA immunization. Each symbol represents an individual mouse serum sample. The geometric mean and standard deviation are shown for each group. The statistical differences between each group were determined and groups with p<0.05 are indicated.</p

    Serum antibody titers against homologous and variant H5N1 <i>ca</i> viruses following different vaccine regimens in ferrets.

    No full text
    <p>Ferrets (N = 4/group) were inoculated with 10<sup>7</sup> PFU of VN04 H5N1 <i>ca</i> intranasally or 200 µg of vector or H5 HA DNA vaccine intramuscularly as indicated. Blood was collected 4 weeks after each dose and H5N1-specific antibody titers in the serum were determined by microneutralization (left panel) and HAI (right panel) assays. Geometric mean titers of neutralizing and HAI Ab against (A) VN04 (A/Vietnam/1203/2004), (B) HK03 (A/Hong Kong/213/2003), (C) AH05 (A/Anhui/1/2005), and (D) IN05 (A/Indonesia/5/2005) <i>ca</i> viruses are depicted. Data points in black indicate titers of post-prime and data points in orange indicate titers of post-boost and the mean Ab titers are indicated in horizontal solid lines. The dashed line indicates the detection limit of the assays.</p

    Schematic design of HBs-L DNA vaccines.

    No full text
    <p>A. These HBs-L DNA vaccines express the full length large protein of the HBsAg (HBs-L) or the truncated large protein with removal of the first 18 amino acids at the N-terminus, respectively. The amino acid positions for each HBs-L immunogen are as indicated. B and C: The ELISA analyses of the HBs-L and HBs-L(T) protein expressions by the relevant DNA and the empty vector as negative control, using commercially available anti-S (B) or anti-Pre-S mAb (C). “S” and “L” represent the supernatant and lysate of transiently transfected 293T cells.</p

    H5 HA DNA vaccine construct.

    No full text
    <p>(A) Schematic diagram of H5-VN.tPA HA gene. The H5-VN.tPA DNA encodes the full length HA antigens with a human tissue plasminogen activator (tPA) leader substituting for the natural HA leader sequence. The cleavage site between HA1 and HA2 subunits are indicated. The numbers above the HA inserts denote the relevant amino acid positions in natural HA proteins. (B) Western-blot analysis of the expression of HA antigen by H5-VN.tPA DNA vaccine in transiently transfected 293T cell supernatant (S) and cell lysate (L).</p

    Protection conferred by different H5 vaccine prime-boost regimens in ferrets.

    No full text
    <p>Four weeks after boost immunization, ferrets were intranasally infected with 10<sup>7</sup> PFU of HK03 <i>ca</i> virus. Nasal turbinates were collected on day 3 post-infection and viral titers in tissue homogenates were determined by EID<sub>50</sub>. Horizontal lines depict the mean virus titers for each group. An asterisk above a horizontal bar denotes statistical significance compared to the vector DNA group (<i>p</i><0.05, Mann-Whitney U Test).</p

    HBs-specific antibody secreting cells (ASC) in splenocytes of immune mice as measured by ELISPOT.

    No full text
    <p>Mice were immunized with HBs-L or HBs-L(T) DNA vaccine, or with empty DNA vector as indicated. The coating antigens were anti-mouse IgG, commercial HBsAg and PBS (Mock) for detection of the total IgG, HBsAg-specific or background level ASC in mouse splenocytes. A: Actual sample wells of HBs-specific ASC spots. B: Frequency of HBs-specific ASC per 10 million splenocytes against different HBsAg coating antigens in each group. Data represent the average of spot forming cells (SFCs) per 10 million of splenocytes from each group of mice plus standard error. The splenocytes were collected 1 week after the last (4<sup>th</sup>) DNA immunization. The statistical differences between groups were determined and difference with p<0.05 are indicated. Each symbol represents an individual mouse serum sample. The geometric mean and standard deviation are shown for each group. The statistical differences between each group were determined and groups with p<0.05 are indicated.</p

    Kinetics of the development of H5 HA-specific serum IgG (A), IgA (B), and neutralizing antibodies (C) elicited by various H5N1 prime-boost regimens.

    No full text
    <p>Ferrets were immunized with either 10<sup>7</sup> PFU of VN04 <i>ca</i> virus (weeks 0 and 4) or 200 µg of H5 HA DNA (weeks 0, 2 and/or 4) and serum samples were collected regularly at 2-week intervals. H5 HA-specific IgG and IgA in ferret serum samples (N = 4/group) were determined by ELISA using recombinant VN04 H5 HA as antigen. Neutralizing antibody titers against 100 TCID<sub>50</sub> of the VN04 <i>ca</i> virus are depicted. Geometric mean titers from each group are shown.</p

    Ferret serum antibody titers from vaccination with iVN04 and VN04 <i>ca</i>.

    No full text
    <p>Ferrets were inoculated with 10<sup>7</sup> PFU of VN04 <i>ca</i> intranasally or 15 µg of iVN04 vaccine intramuscularly. Blood was collected 4 weeks after each dose and the H5N1-specific antibody levels (expressed as geometric mean titers) were determined by microneutralization assay.</p
    corecore