mouse ifn γ elispot kit  (R&D Systems)

Journal: Molecular Therapy Oncology

Article Title: Induced pluripotent stem cell-derived dendritic cells potentiate the efficacy of interleukin-12-expressing oncolytic HSV-1

doi: 10.1016/j.omton.2026.201252

Figure Lengend Snippet: Immunological assessments of the combination of T-mfIL12 and immature BMDCs (A) Immunohistochemical analysis of representative MB49 tumors from the indicated treatment groups. Increased intratumoral infiltration of CD8 + lymphocytes was most prominent in the T-mfIL12+BMDC group, followed by the T-01+BMDC group. In both groups, elevated levels of CD4 + lymphocyte infiltration were also observed in the same tumor sections. Other groups, except for the mock group, showed moderate levels of CD4 + infiltration. Scale bars: 100 μm. (B) ELISpot assay for IFN-γ expression revealed the highest number of IFN-γ-producing splenocytes in the T-mfIL12+BMDC group, followed by the T-01+BMDC group. Monotherapies with BMDC, T-01, or T-mfIL12, as well as the mock group, did not result in significant increases. n = 3 per group; all samples were analyzed in triplicate. Error bars represent standard deviation (SD). ∗ p < 0.05, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: Cells were plated in triplicates at a density of 1 × 10 5 cells per well in 50 μL of RPMI-1640 medium supplemented with 10% FCS, 2% penicillin-streptomycin, and 50 μM 2-ME, using a MultiScreen 96-well plate (Millipore, USA) pre-coated with anti-mouse IFN-γ monoclonal antibodies from the Mouse IFN-γ ELISpot Kit (AN18; Mabtech AB, cat. #3321-2A).

Techniques: Immunohistochemical staining, Enzyme-linked Immunospot, Expressing, Standard Deviation

Journal: Molecular Therapy Oncology

Article Title: Induced pluripotent stem cell-derived dendritic cells potentiate the efficacy of interleukin-12-expressing oncolytic HSV-1

doi: 10.1016/j.omton.2026.201252

Figure Lengend Snippet: Immunological assessments of the combination of T-mfIL12 and immature iPSDCs (A) Immunohistochemical analysis of representative MB49 tumors from the indicated treatment groups. Increased intratumoral infiltration of CD8 + lymphocytes was observed in both the T-mfIL12+BMDC and T-mfIL12+iPSDC groups. In both groups, CD4 + lymphocyte infiltration was also more prominently observed in the same tumor sections. Other treatment groups, excluding the mock group, exhibited moderate levels of CD4 + T cell infiltration. Scale bars: 100 μm. (B) ELISpot assay showed a high and comparable increase in IFN-γ-producing splenocytes in both the T-mfIL12+BMDC and T-mfIL12+iPSDC groups. n = 3 mice per group; all samples were analyzed in triplicate. Error bars represent standard deviation (SD). ∗∗∗∗ p < 0.0001; NS, not significant.

Article Snippet: Cells were plated in triplicates at a density of 1 × 10 5 cells per well in 50 μL of RPMI-1640 medium supplemented with 10% FCS, 2% penicillin-streptomycin, and 50 μM 2-ME, using a MultiScreen 96-well plate (Millipore, USA) pre-coated with anti-mouse IFN-γ monoclonal antibodies from the Mouse IFN-γ ELISpot Kit (AN18; Mabtech AB, cat. #3321-2A).

Techniques: Immunohistochemical staining, Enzyme-linked Immunospot, Standard Deviation

Journal: Molecular Therapy. Nucleic Acids

Article Title: Comparative analysis of expression, immunogenicity, and safety profiles between linear and circular RNA vaccine platforms

doi: 10.1016/j.omtn.2026.102954

Figure Lengend Snippet: Cell-mediated immune responses to different RNA platforms Each group includes a negative control (DPBS), unmodified linear mRNA (Lin-WT), modified linear mRNA (Lin-m1Ψ), and unmodified circular RNA (Circ-WT). (A) Immunization schedule schematic diagram for assessing T cell responses. Balb/c mice were intramuscularly immunized twice, 2 weeks apart, with 10 μg of LNP-encapsulated HA-encoding mRNA (Lin-WT, Lin-m1Ψ, or Circ-WT). (B and C) IFN-γ ELISPOT assay showing the number of antigen-specific IFN-γ-secreting splenocytes following immunization with HA-encoding mRNA. (D–F) Flow cytometry analysis of cytokine-producing CD8+ T cells. The frequencies of IFN-γ+, IL-2+, and TNF-α + CD8+ T cells were assessed to evaluate antigen-specific T cell activation. (G–I) Cytokine-producing CD4+ T cells, with frequencies of IFN-γ+, IL-2+, and TNF-α+ CD4+ T cells measured by intracellular cytokine staining. (J) Analysis of double-positive cytokine-expressing CD8+ T cells, indicating polyfunctional T cell responses. (K) Analysis of double-positive cytokine-expressing CD4+ T cells, indicating helper T cell activation. Data represent mean ± SD ( n = 5 per group). Statistical significance was determined by ordinary one-way ANOVA with Tukey’s test or the Kruskal-Wallis test with Dunn’s multiple comparison test, depending on the normality of the data. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗∗ p ≤ 0.0001.

Article Snippet: IFN-γ secreting T cells were detected using the ELISpot assay and the mouse IFN-γ ELISpot BASIC kit from Mabtech (Stockholm, Sweden), following the manufacturer’s instructions.

Techniques: Negative Control, Modification, Enzyme-linked Immunospot, Flow Cytometry, Activation Assay, Staining, Expressing, Comparison

Journal: Molecular Therapy. Nucleic Acids

Article Title: Comparative analysis of expression, immunogenicity, and safety profiles between linear and circular RNA vaccine platforms

doi: 10.1016/j.omtn.2026.102954

Figure Lengend Snippet: Differential innate immune responses induced by different RNA platforms Groups comprise the negative control (DPBS), unmodified linear mRNA (Lin-WT), modified linear mRNA (Lin-m1Ψ), and unmodified circular RNA (Circ-WT). (A) Schematic presentation of in vivo cytokine analysis. Mice were injected intramuscularly with LNP-encapsulated mRNA, and cytokine levels were assessed in serum and lymph nodes at 6- and 24-h post-inoculation. (B–G) In vivo cytokine responses measured in serum and lymph nodes at different time points. Levels of (B-C) IFN-γ, (D) RIG-I, (E) IFN-β, (F) TNF-α, (G) IL-6. mRNA Fold change calculated by the 2 −ΔΔCt method and normalized to GAPDH, with values expressed as fold change relative to the DPBS group. Data represent mean ± SD ( n = 5 per group). Statistical significance was determined by ordinary one-way ANOVA with Tukey’s test or the Kruskal-Wallis test with Dunn’s multiple comparison test, depending on the normality of the data. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001, ∗∗∗∗ p ≤ 0.0001.

Article Snippet: IFN-γ secreting T cells were detected using the ELISpot assay and the mouse IFN-γ ELISpot BASIC kit from Mabtech (Stockholm, Sweden), following the manufacturer’s instructions.

Techniques: Negative Control, Modification, In Vivo, Injection, Comparison

Journal: bioRxiv

Article Title: Coordinated Tuning of Ionizable Lipids and Formulation Redirects mRNA Vaccines Toward Lymphoid-Specific CD4 + T Cell Immunity

doi: 10.64898/2026.04.16.719092

Figure Lengend Snippet: (a) Vaccination scheme. BALB/c mice were immunized intramuscularly with SARS-CoV-2 spike mRNA-loaded LNPs (0.25 mg/kg) following a prime–boost regimen (day 0 and day 21). Blood samples and splenocytes were collected three weeks after the booster dose for antibody and T cell analysis. (b) Antigen-specific IgG levels at three weeks post-boost (n = 5 per group). (c) Neutralizing antibody titers assessed by PRNT₅₀ at three weeks post-boost (n = 3 per group). (d) Antigen-specific T cell responses assessed by IFN-γ ELISpot (n= 2–3 per group). (e) Viral challenge scheme. K18-hACE2 transgenic mice were immunized following the same prime–boost regimen and challenged with SARS-CoV-2 three weeks after the booster dose. (f) Body weight changes following viral challenge (n = 5 per group). (g) Survival curves following viral challenge (n = 5 per group). Statistical significance for (b–d) was determined by one-way ANOVA followed by Tukey’s multiple comparisons test. Survival curves were compared using the log-rank (Mantel-Cox) test. Data are presented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: The assay was performed using a mouse IFN-γ ELISpot kit (XEL485, R&D Systems) according to the manufacturer’s instructions.

Techniques: Cell Analysis, Enzyme-linked Immunospot, Transgenic Assay

Journal: eBioMedicine

Article Title: Nasal RNA-scaffold-protein vaccine protects mice from human H5N1 clade 2.3.4.4b virus lethal infection and safeguards against vaccine-unmatched viruses

doi: 10.1016/j.ebiom.2026.106228

Figure Lengend Snippet: Mutant proteins impacted RSPVac immunisation. (A) Schematic showing the disrupted formation of RSPVac using an RNA-binding-deficient mutant. Female BALF/c mice (n = 6) were nasally immunised with two doses of RSPVac-H5, 14-days apart. The RSPVac was produced with either the WT or mutant protein. Sera, BALF, and BALF-flushed cells were harvested 10 days post-2nd dose and analysed. (B) Fluorescence polarisation of WT and Mutant protein. After mutating major arginine residues, the RNA-binding affinity was reduced by 7-fold. (C–F) ELISA for anti-HA serum IgG, BALF IgA, Serum IgG subtypes, and BALF IgG subtypes. (G) IFNγ ELISpot analysis of BALF-flushed cells, stimulated with the immunogen (WT protein used to generate RSPVac-H5-1194). (A) Schematic of the phase separation mutant of the SARS2-RSPVac. To explore whether phase separation is related to RSPVac nasal immunisation, phase separation mutants (Psmut) were generated and used to generate SARS2-RSPVac. Female BALB/c mice were immunised using the same regimen with WT or Psmut RSPVac. Sera and BALF were harvested for downstream analysis. (B–F) ELISA for anti-spike RBD serum IgG, BALF IgG, BALF IgA, serum IgG1/IgG2a, BALF IgG1/IgG2a. (G) IFNγ ELISpot analysis of BALF-flushed cells, stimulated with the immunogen (protein used to generate WT SARS2-RSPVac). Statistical significance was determined by the Mann–Whitney test. p < 0.05 was considered significant. P-values were shown, and those that were considered statistically not significant were labelled not significant (ns). Figures A were created in BioRender.

Article Snippet: BALF cells were stimulated with recombinant proteins and IFN-secreting cells were detected by ELISpot Flex Mouse IFN-γ (ALP) kit (Mabtech Cat# 3321-2A).

Techniques: Mutagenesis, RNA Binding Assay, Produced, Fluorescence, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, Generated, MANN-WHITNEY

Journal: eBioMedicine

Article Title: Nasal RNA-scaffold-protein vaccine protects mice from human H5N1 clade 2.3.4.4b virus lethal infection and safeguards against vaccine-unmatched viruses

doi: 10.1016/j.ebiom.2026.106228

Figure Lengend Snippet: Mouse antibody and T cell responses following RSPVac nasal vaccination. (A) Vaccination regimen. Female BALB/c mice (n = 6–9) were intranasally vaccinated with 2 doses of RSPVac, 14 days apart. Sera, BALF, and BALF cells were harvested at day 10 post-2nd dose for analysis. Control mice received the protein component without RNA (protein-only). Data was shown as box plots with respective colours indicating vaccine given, showing all values. Error bars represented the highest and lowest values of each box. ELISA was used to measure antibody responses for (B) anti-HA serum IgG, (C) anti-HA BALF IgG, (D) anti-HA BALF IgA, (E) anti-NP, and (F) serum anti-HA IgG subtypes for each of the RSPVac, respectively. (G) Mucosal T cell responses measured by IFNγ+ ELISpot using the immunogen (the protein used to generate RSPVac) stimulation of BALF cells. (H) To analyse lung T cells, female BALB/c mice (n = 6) were intranasally vaccinated with 2 doses of RSPVac, 14 days apart. Lungs were harvested at day 7 post-2nd dose. Single cell suspensions were prepared, and the NP peptide pool of H5N1, H1N1, and H7N9 was used to stimulate lung cells. Resident T cell responses were analysed by intracellular staining and FACS analysis. NP-reactive (I) CD4+ and (J) CD8+ T cells were shown, comparing protein-only and RSPVac-vaccinated. Statistical significance was determined by the Mann–Whitney test. p < 0.05 was considered significant. P-values were shown, and those that were considered insignificant were labelled not significant (ns). Figures A and H were created in BioRender.

Article Snippet: BALF cells were stimulated with recombinant proteins and IFN-secreting cells were detected by ELISpot Flex Mouse IFN-γ (ALP) kit (Mabtech Cat# 3321-2A).

Techniques: Control, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, Single Cell, Staining, MANN-WHITNEY