PMC

Figure 3. Anti-HER2/neu T-cell immune response after vaccination as detected by HER2/neu-p63-specific pentamer staining. Splenocytes from wild-type (WT) BALB/c mice intradermally immunized with pDNA(HER2/neu) using gene gun (GG) or jet injector (JI) were stained with HER2/neu-p63-specific pentamers. For cytofluoromtetric analyses, splenocytes from mice belonging to the same experimental group were pooled. Splenocytes were analyzed within a CD3⁺-restricted gate.

Figure 5. Humoral anti-HER2/neu immune responses after vaccination. (A-D) Antibody responses (total IgG and IgG isotypes) against HER2/neu were determined by a cytofluorometric assay in wild-type (WT) BALB/c mice intradermally immunized with pDNA(HER2/neu) using gene gun (GG) or jet injector (JI). Mice had been immunized with DNA on days 1 and 15. Cytofluorometric assays were performed 7 d after the last vaccination. (A) Anti-HER2/neu, total IgG. (B) Anti-HER2/neu, IgG1. (C) Anti-HER2/neu, IgG2a. (D) Anti-HER2/neu, IgG2b. * = statistically significant vs. all other groups (p < 0.05).

Figure 2. Anti-HER2/neu T-cell immune responses after vaccination. (A, B) Splenocytes from wild-type (WT) BALB/c mice intradermally immunized with pDNA(HER2/neu) using gene gun (GG) or jet injector (JI) were stimulated with different peptide combinations (derived from the extracellular domain or the intracellular domain of HER2/neu, or both). Specific T-cell responses were analyzed by a interferon γ (IFNγ)-specific (A) or interleukin-4 (IL-4)-specific ELISpot assays. Mice had been immunized with DNA on days 1 and 15. ELISpot assays were performed 7 d after the last vaccination. For ELISpot assays, splenocytes within the different groups of mice were pooled. ECD, extracellular domain; ICD, intracellular domain. * = statistically significant vs. all control groups (p < 0.05).

Figure 4. CTL assays after vaccination with gene gun or jet injector. (A-C) Wild-type (WT) BALB/c mice were immunized by gene gun (GG) or jet injector immunization (JI) with pDNA(HER2/neu), mock vector (pVax) or gold particles/PBS on days 1 and 15. On day 22, splenocytes were restimulated in vitro for 5 d with irradiated BALB/c 3T3 cells that were pulsed with a HER2/neu-p63 peptide. CTL activity was measured in a standard ⁵¹Cr release assay using HER2/neu⁺ D2F2/E2 tumor cells (A), HER2/neu^- D2F2 tumor cells, pulsed with the HER2/neu-p63 peptide (B), or HER2/neu^- D2F2 tumor cells only (negative control) (C). * = statistically significant vs. all other groups (p < 0.05).

Figure 1. Short- and long-term tumor protection by intradermal DNA vaccination using gene gun or jet injector: percentage of tumor-free mice after vaccination and tumor challenge. (A, B) Wild-type (WT) BALB/c mice were immunized by gene gun (GG) or jet injector (JI) delivery with pDNA(HER2/neu), mock vector (pVax) or gold particles/PBS on days 1 and 15. On day 25 tumor challenge was performed with 2 × 10⁵ HER2/neu⁺ syngeneic D2F2/E2 tumor cells. Tumor growth was then monitored thereafter until day 140. n = 10 for each group of mice. (A) Short-term protection (day 50). (B) Long-term protection (day 140). * = statistically significant vs. all other groups (p < 0.05)