Journal: Vaccine

Article Title: Immunogenic potential of DNA vaccine candidate, ZyCoV-D against SARS-CoV-2 in animal models

doi: 10.1016/j.vaccine.2021.05.098

Figure Lengend Snippet: Detection of IFN-γ responses in BALB/c mice post-administration of DNA vaccine. BALB/c mice were immunized with 25 and 100 μg of DNA vaccine. IFN-γ responses were analyzed in the animals on days 14, 28 and 42. T cell responses were measured by IFN-γ ELISpot in splenocytes stimulated for 24 h with overlapping peptide pools spanning the SARS-CoV-2 spike region.

Article Snippet: The 96-well Mouse IFN-γ ELISPOT kit (CTL, USA) plates pre-coated with purified anti-mouse IFN-γ capture antibody were taken out blocked with RPMI-1640 + 10% FBS + 1X Antibiotic-Antimycotic for 1 Hr in CO 2 incubator.

Techniques: Enzyme-linked Immunospot

Journal: Microbial biotechnology

Article Title: Rectal single dose immunization of mice with Escherichia coli O157:H7 bacterial ghosts induces efficient humoral and cellular immune responses and protects against the lethal heterologous challenge

doi: 10.1111/j.1751-7915.2011.00316.x

Figure Lengend Snippet: Cellular immune responses stimulated in mice after the immunizations with EHEC O157:H7 (N°CIP 105282) BGs. Mice were immunized as described in Fig. 1 . Spleen cells (1 × 10 5 per well) were incubated for 16 h in the presence of EHEC O157:H7 BGs (specific Ag); measles inactivated Ag (strain Edmonston, non‐specific Ag) or as a negative control spleen cells were left without stimulation. The numbers of IFN‐γ‐producing cells were determined by ELISPOT assays. Rectal immunization with EHEC O157:H7 BGs effectively increased the number of IFN‐γ‐producing spleen cells after recognition of specific Ag (A). The stimulatory effect of the boost immunization was detected on day 42 (14 days after the second immunization) and remained significant until the end of experiment (B). Data represent the mean of three independent experiments ± SD (three mice per time point). P ‐values < 0.05 were considered significant and are indicated with asterisks (* P < 0.05; ** P < 0.01; *** P < 0.001). a, day of the first immunization in all groups; b, day of the second immunization in groups B1 and D1; c, day of the challenge.

Article Snippet: The number of IFN‐γ‐producing cells was determined by Mouse IFN‐γ ELISpot Kit (R&D Systems, Inc, Minneapolis, USA) following the manufacturer's instructions.

Techniques: Incubation, Negative Control, Enzyme-linked Immunospot

Journal: Oncotarget

Article Title: The novel complex combination of alum, CpG ODN and HH2 as adjuvant in cancer vaccine effectively suppresses tumor growth in vivo

doi: 10.18632/oncotarget.17504

Figure Lengend Snippet: ( A ) The 4-h NK assay against YAC-1 targets was determined at various effector:target ratios. Error bars represent mean + SEM. ** p < 0.01, *** p < 0.001. ( B ) The cytolytic activity of splenocytes against NY-ESO-1 + B16 cells was measured using a 4 h 51 Cr-release assay at various E:T ratios. Error bars represent mean + SEM. *** p < 0.001. ( C ) The representative graphs of spot-forming cells for IFN-γ and IL-4 at one week post-immunization in the ELISpot assay. ( D ) The average number of IL-4/IFN-γ secreting splenocytes was calculated (n = 3, three independent experiments). Error bars represent mean + SEM. * p < 0.05, ** p < 0. 01. ( E ) T cell analysis in splenocytes after vaccination. Intracellular staining of IFN-γ in CD4 + and CD8 + T cells was analyzed by FACS.

Article Snippet: The mouse IFN-γ/IL-4 Dual-Color ELISpot kit (R&D Systems, Minneapolis, USA) was used for ELISpot assay.

Techniques: Activity Assay, Release Assay, Enzyme-linked Immunospot, Cell Analysis, Staining

Journal: Journal of Nanobiotechnology

Article Title: Double-layered N-S1 protein nanoparticle immunization elicits robust cellular immune and broad antibody responses against SARS-CoV-2

doi: 10.1186/s12951-024-02293-y

Figure Lengend Snippet: Evaluation of splenic cellular immune responses in immunized mice. A Enumeration of Interferon-gamma (IFN-γ) and Interleukin-4 (IL-4) secreting splenocytes clones re-stimulated with N peptide pool or S1 protein were determined by ELISpot assay. B The levels of IFN-γ, IL-4, and IL-2 secreted in the splenocytes culture medium from immunized mice were detected by cytokine ELISA after stimulation for 2.5 days. C The SI of lymphocyte proliferation assay. D–G Splenocytes were analyzed in FCM. Percentage of activated CD4 + ( E ) and CD8 + ( G ) T cells. D , F Representative pseudocolor plots (smooth on). Bars are presented as mean ± SEM of triplicate assays, where n = 3 mice in each group. Individual values are indicated by colored symbols. Statistical significance was assessed by one-way ANOVA analysis followed by multiple comparisons post hoc tests in A-B, and corresponding P values shown in bar charts. ***p-value < 0.001, **p-value < 0.01, *p-value < 0.05, ns p-value > 0.05

Article Snippet: On day 21st after the boost immunization, splenocytes of vaccinated mice were collected from all immunization groups as single-cell suspensions using the mouse spleen lymphocyte isolation kit (Solarbio; P8860).

Techniques: Clone Assay, Enzyme-linked Immunospot, Enzyme-linked Immunosorbent Assay, Lymphocyte Proliferation Assay

Journal: Molecular Therapy. Nucleic Acids

Article Title: A single-chain mRNA vaccine co-expressing GPC and NP provides complete protection against lethal Dabie bandavirus challenge in mice

doi: 10.1016/j.omtn.2025.102755

Figure Lengend Snippet: Cellular immune responses elicited by BDV mRNA vaccines in immunocompetent C57BL/6J mice (A and B) T cell responses to BDV antigens were analyzed via an ELISpot assay. Splenocytes were collected 10 days after the second immunization, and IFN-γ-, TNF-α-, IL-2-, and IL-4-secreting cells were generated in response to the SFTSV GP and NP peptides. (C and D) Flow cytometry analysis of IFN-γ in SFTSV-antigen-specific CD4 + T and CD8 + T cells and central memory CD8 + T cells (Tcm) in the spleens of immunized mice. Each dot represents serum from one mouse. The experiments were conducted independently. The data are presented as the mean ± SEM. Statistical significance was assessed via one-way ANOVA with multiple comparisons tests (∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001).

Article Snippet: The secretion of the antigen-specific IFN-γ, TNF-α (Mebtech, Bristol, USA), IL-2, and IL-4 by splenocytes was measured via mouse ELISpot kits (Dakewe, Shenzhen, China) according to the manufacturer’s instructions.

Techniques: Vaccines, Enzyme-linked Immunospot, Generated, Flow Cytometry

Journal: NPJ Vaccines

Article Title: Shigella virulence protein VirG is a broadly protective antigen and vaccine candidate

doi: 10.1038/s41541-023-00797-6

Figure Lengend Snippet: a , b Spleen and bone marrow were obtained from mice immunized IM with 20 µg of VirGα (IM) or IN with S. flexneri 2a (sl Sf2a-IN) that survived S. flexneri 2a lethal challenge. Single-cell suspensions from individual mice in each group were pooled, and VirGα-specific and total IgG and IgA ASC were measured by ELISpot in quadruplicates. Bars represent mean values per 10 6 cells ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001 by t test.

Article Snippet: The frequencies of total and VirGα-specific IgG- and IgA-secreting cells were determined using a mouse IgA/IgG double-color ELISpot assay kit [Catalogue # mIgGIgA-DCE-1M/2, ImmunoSpot® ELISpot Kit, Cellular Technology Limited (CTL), Cleveland, OH] according to the manufacturer’s instructions.

Techniques: Enzyme-linked Immunospot

Journal: Vaccine

Article Title: Adjuvants that stimulate TLR3 or NLPR3 pathways enhance the efficiency of influenza virus-like particle vaccines in aged mice

doi: 10.1016/j.vaccine.2011.09.051

Figure Lengend Snippet: Adult and aged mice (n=5) were challenged with a sub-lethal dose of the Indo/05-PR8 virus and (A) the virus titer in the lung and (B) the IFNα levels in the lungs at day 3, 6 and 9 post infection were determined (mean ± SEM). Additionally aged and adult mice were infected for 10 days and (C) the percentage of the HA533 and NP147 epitope specific CD8+ T cells was determined by pentamer staining and in (D) the number of HA533-pentamer and NP147-pentamer positive CD8+ T cells is shown. The whiskers in (C) and (D) designate the minimum and maximum values. The bar within the box designates the median. (E) IFNγ ELISPOT analysis with splenocytes collected 10 days post infection. * indicates p<0.05.

Article Snippet: IFNα levels in lung homogenates were quantified using a commercial mouse IFNα ELISA kit (PBL Biomedical Laboratories, Piscataway, NJ) according to the manufacturer instructions.

Techniques: Virus, Infection, Staining, Enzyme-linked Immunospot

Journal: Vaccine

Article Title: Adjuvants that stimulate TLR3 or NLPR3 pathways enhance the efficiency of influenza virus-like particle vaccines in aged mice

doi: 10.1016/j.vaccine.2011.09.051

Figure Lengend Snippet: Groups of mice (n=8-10) were immunized with H5N1 Indo/05 VLP vaccine in combination with Imject® Alum via the intra-muscular route. Antibodies elicited to the VLP vaccine with or without Imject® Alum were determined by (A) hemagglutination inhibition assay and (B) ELISA after the second immunization (week 5, ELISA compares VLP immunization in combination with Imject® Alum in both age groups) (±SEM). (C) Mice were challenged with a lethal dose of Indo/05-PR8 virus and monitored daily for weight loss and mortality. The data are plotted as percentage of the initial body weight. Statistical analysis was performed by determining the p-value from comparison of percentage of initial body weight of aged vs. adult mice receiving the VLP + ImjectAlum and comparison of the percentage of initial body weight of the naïve aged vs. adult mice at the respective time points. (D) Separate groups of VLP+Imject® Alum immunized and naïve mice of both age groups were sacrificed on day 6 post infection and HA533-pentamer staining was performed. The percentage of HA533 pentamer + CD8+ T cells is shown for both age groups. The whiskers in (D) designate the minimum and maximum values. The bar within the box designates the median. * indicates p<0.05.

Article Snippet: IFNα levels in lung homogenates were quantified using a commercial mouse IFNα ELISA kit (PBL Biomedical Laboratories, Piscataway, NJ) according to the manufacturer instructions.

Techniques: HI Assay, Enzyme-linked Immunosorbent Assay, Virus, Comparison, Infection, Staining

Journal: Vaccine

Article Title: Adjuvants that stimulate TLR3 or NLPR3 pathways enhance the efficiency of influenza virus-like particle vaccines in aged mice

doi: 10.1016/j.vaccine.2011.09.051

Figure Lengend Snippet: Mice (n=6-8) were immunized with H5N1 Indo/05 VLP vaccine alone or in combination with poly (I:C) via the intranasal route. As controls, mice were immunized with poly (I:C) alone or were non-immunized. (A) Antibodies elicited to the vaccine were determined by ELISA and (B) hemagglutination inhibition assay. In (C) aged and (D) adult mice were challenged with a lethal dose Indo/05-PR8 and monitored daily for weight loss and mortality. The data are plotted as percentage of the initial body weight. Statistical analysis was performed by comparing the percentage of initial body weight of the group receiving the VLP alone to the group receiving the VLP in combination with poly (I:C). (E) Mice immunized with the VLP vaccine alone were sacrificed 6 days post infection (DPI) and the number of HA-specific IgG and IgA antibody secreting cells in the lungs was determined.. * indicates p<0.05.

Article Snippet: IFNα levels in lung homogenates were quantified using a commercial mouse IFNα ELISA kit (PBL Biomedical Laboratories, Piscataway, NJ) according to the manufacturer instructions.

Techniques: Enzyme-linked Immunosorbent Assay, HI Assay, Infection