Journal: Molecular Therapy Advances

Article Title: Efficacy and safety of SENS-501, a dual-AAV otoferlin gene therapy, for DFNB9 congenital deafness

doi: 10.1016/j.omta.2026.201762

Figure Lengend Snippet: Intracochlear injection of SENS-501 in NHPs results in a mild humoral and an undetectable cellular response to the capsid (A) AAV8 anti-drug antibody (ADA) titers measured in serum at pre-dose and at 16, 29, and 92 days post-vector administration in NHPs. (B) Anti-AAV8 neutralizing antibodies (NAb) titers measured in serum at pre-dose and at 16, 29, and 92 days post-vector administration in NHPs. (C and D) IFN-γ spot forming units (SFUs) measured by ELISpot assay at 29 (C; left) and 92 (D; right) days post-injection. Peripheral blood mononuclear cells (PBMCs) from the indicated groups were stimulated with three different AAV8 peptide pools and a positive control (PMA/ionomycin). The dotted line represents the assay-specific positivity threshold. Each dot represents one animal. Bars represent the mean ± SEM.

Article Snippet: After the incubation, detection was performed with a monoclonal anti-monkey IFN-γ antibody (Monkey IFN-γ ELISpot Pro Kit, Mabtech) coupled with alkaline phosphatase and incubated with BCIP/NBT (5-bromo-4-chloro-3-indolyl-1-phosphate / nitroblue tetrazolium) substrate to detect secreted IFN-γ.

Techniques: Injection, Plasmid Preparation, Enzyme-linked Immunospot, Positive Control

Journal: Poultry Science

Article Title: A novel self-amplified RNA vaccine co-expressing NA and HA1 delivered by Salmonella confers potent protection against H9N2 influenza in chickens

doi: 10.1016/j.psj.2026.107072

Figure Lengend Snippet: Intracellular cytokine production. Chicken splenic lymphocytes were isolated for analysis. Cell proliferation was assessed using CCK-8 analysis with ConA (A), mixed HA1 peptides (B), and NA protein (C). Additionally, the production of IFN-γ by splenic T lymphocytes was measured via an ELISpot assay, utilizing NA and HA1 proteins as stimulators for 36 h (D).

Article Snippet: IFN-γ production was assessed using a commercial Chicken IFN-γ ELISpot kit (Mabtech, Sweden).

Techniques: Isolation, CCK-8 Assay, Enzyme-linked Immunospot

Journal: Poultry Science

Article Title: A novel self-amplified RNA vaccine co-expressing NA and HA1 delivered by Salmonella confers potent protection against H9N2 influenza in chickens

doi: 10.1016/j.psj.2026.107072

Figure Lengend Snippet: Intracellular cytokine production. The intracellular mRNA expression levels of IL-4 (B, D) and IFN-γ (A, C)—as well as the relative concentrations of these cytokines in cell culture supernatants stimulated by the NA peptide (E, F) or HA1 protein (G, H) for 48 h—were determined using qRT-PCR and ELISA, respectively. Data are expressed as the mean ± SEM and analyzed using one-way ANOVA (* P < 0.05, ** P < 0.01, and *** P < 0.001; n = 4).

Article Snippet: IFN-γ production was assessed using a commercial Chicken IFN-γ ELISpot kit (Mabtech, Sweden).

Techniques: Expressing, Cell Culture, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: Bioactive Materials

Article Title: Splenic dendritic cell-targeting mRNA transfection of H-type ionizable lipid-based LNPs for enhancing tumor immunotherapy

doi: 10.1016/j.bioactmat.2026.02.018

Figure Lengend Snippet: In vivo DC maturation and T cell activation in C57BL/6J mice induced by mOVA/H 18 NPs through intravenous injection. C57BL/6J mice were vaccinated with different formulations on Day 0 and Day 5. On Day 10, the spleens of mice were collected and analyzed by flow cytometry. Quantification analysis of (A) CD80 + CD86 + DCs and (B) CD40 + DCs in the spleen. Quantification analysis of (C) CD3 + CD4 + T cells and (D) CD3 + CD8 + T cells in the spleen. (E) Quantification analysis and (F) representative flow cytometry contour plots of OVA-specific CD8 + T cells among all cell populations in the spleen. (G) Quantification analysis and (H) representative flow cytometry contour plots of IFN-γ + CD8 + T cells among all cell populations in the spleen. (I) Quantification results and (J) representative images of IFN- γ -secreting immune cells in the spleen of mice analyzed by enzyme-linked immunospot (ELISpot) assay. Data were shown as mean ± SD (n = 3).

Article Snippet: Quantification analysis of (C) CD3 + CD4 + T cells and (D) CD3 + CD8 + T cells in the spleen. (E) Quantification analysis and (F) representative flow cytometry contour plots of OVA-specific CD8 + T cells among all cell populations in the spleen. (G) Quantification analysis and (H) representative flow cytometry contour plots of IFN-γ + CD8 + T cells among all cell populations in the spleen. (I) Quantification results and (J) representative images of IFN- γ -secreting immune cells in the spleen of mice analyzed by enzyme-linked immunospot (ELISpot) assay.

Techniques: In Vivo, Activation Assay, Injection, Flow Cytometry, Enzyme-linked Immunospot

Journal: iScience

Article Title: Integrating complementary approaches reveals antigen-reactive CD4 + T cell states after SARS-CoV-2 vaccination

doi: 10.1016/j.isci.2026.116175

Figure Lengend Snippet: Identification of SARS-CoV-2-specific T cell responses by reverse phenotyping (A) CoVa-Adapt study design and sample collection scheme. For all donors, PBMCs were collected at day 0 (P0), 10 days after primary (P10), 10 and 210 days after secondary (S10, S210), and 10 and 189 days after tertiary (T10, T189) vaccination. For selected donors, PBMCs were additionally sampled 108 days after tertiary vaccination (T108, n = 7). Vaccination-induced T cell responses were characterized for most donors on a quantitative level by IFNγ ELISpot. Selected CoVa-Adapt donors were subjected to in-depth characterization using scRNAseq (reverse phenotyping and epitope-specific analyses) followed by TCR functional testing. (B–G) scRNAseq data from the reverse phenotyping dataset. For reverse phenotyping, PBMCs were re-stimulated with 15-mer peptides covering the complete wild-type spike protein or left untreated. Sorted non-naïve CD4 + and/or CD8 + T cells were subjected to scRNAseq. Only CD4 + T cells are shown (annotation described in the methods section). The full dataset is depicted in . (B) UMAP of stimulated (blue) and unstimulated (orange) T cells (left) and Leiden clusters (right; cluster names in UMAP, cluster numbers on the right) ( n = 101,939 cells in total). Cells located within the reactive cluster are displayed with increased point size. (C) Dot plots of log-normalized expression of representative genes per cluster. Selected genes of the reactive cluster are highlighted in gray. Numbers on the left indicate cluster numbers with reactive cluster 12 highlighted in bold. (D and E) IFNG expression (D) and proliferation score (E) in unstimulated (stimulated cells in gray) and stimulated (unstimulated cells in gray) CD4 + T cells (left), and quantification in the stimulated condition of cells in the reactive cluster versus all other clusters (right). Cells located within the reactive cluster are displayed with increased point size. For IFNG , cells with log-normalized gene expression of 0 are shown in gray in UMAPs. Statistical testing by the Mann-Whitney U test. (F) UMAP visualization of cells classified as reactive (cells located in the reactive cluster or belonging to clones where at least one cell is in the reactive cluster) from donor A5 at individual time points after primary (P), secondary (S), and tertiary (T) vaccination in the stimulated condition. Color gradient indicates IFNG expression at the indicated time points. Non-reactive cells and cells from other donors are shown in gray. (G) Fraction of cells from donor A5 at each time point belonging to the reactive cluster. (H and I) Identification of spike-reactive T cells after 20h of in vitro re-stimulation of PBMCs with 15-mer peptides covering the complete wild-type spike protein. Peptides were provided in two subpools, S1 (depicted in H) and S2. Primary data (H) of donor A5 is shown. Quantification (I) of spot-forming units (SFU) for IFNγ ELISpot (combined frequencies of S1 and S2 subpools), data points represent individual donors ( n = 12–19 per time point), solid lines indicate the mean. Samples without SFU above the negative control were set to not detected (n.d.). Donor A5 is highlighted in pink. Statistical testing by the Kruskal-Wallis test followed by Dunn’s multiple comparisons test. Significant differences from the P10 time-point are indicated. ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001, n.s. not significant.

Article Snippet: Plates were washed with PBS containing 0.05% Tween 20 (Sigma-Aldrich, P9416-50 mL) and incubated with biotinylated anti-human IFNγ monoclonal antibody (clone 7-B6-1, Mabtech, 3420-6-250) at 0.2 μg/well for 2 h. Plates were washed a second time with PBS containing 0.05% Tween 20 and subsequently incubated with an avidin-biotinylated peroxidase complex (VECTASTAIN Elite ABC-HRP Kit, Vector Laboratories, VEC-PK-6100) for 1–2 h. Afterward, plates were washed first with PBS containing 0.05% Tween 20 following one washing step with PBS.

Techniques: Enzyme-linked Immunospot, Functional Assay, Expressing, Gene Expression, MANN-WHITNEY, Clone Assay, In Vitro, Negative Control

Journal: iScience

Article Title: Distinct CD8 + T cell types associated with COVID-19 severity in unvaccinated HLA-A2 + patients

doi: 10.1016/j.isci.2026.115880

Figure Lengend Snippet: Information on 11 selected HLA-A2-binding 9-mer peptides derived from the SARS-CoV-2 genome (A) Workflow of HLA-A2-binding peptide prediction with NETMHCpan4.1 and IFN-γ ELISpot assay of PBMCs from 24 patients with COVID-19 using 31 predicted peptides ( and ). (B) Homology of 11 selected peptides among SARS-CoV-2 variants and other strains (as indicated). “YES” indicates that more than 90% of epitope sequences matched within 200 selected peptide sequences from the database . Otherwise, the % of matched sequences is shown. “NO” indicates no matched sequences. (C) Information on the SARS-CoV-2 genome and mapping of 11 selected peptides on its genome.

Article Snippet: anti-human-IFN-γ antibody , MABTECH , Cat# 3420-3-250; RRID: AB_907283.

Techniques: Binding Assay, Derivative Assay, Enzyme-linked Immunospot

Journal: iScience

Article Title: Distinct CD8 + T cell types associated with COVID-19 severity in unvaccinated HLA-A2 + patients

doi: 10.1016/j.isci.2026.115880

Figure Lengend Snippet: IFN-γ ELISpot of CD8 + T cells in PBMCs from HLA-A2 + patients with COVID-19 using selected SARS-CoV-2 peptides (A) Cross-sectional ELISpot assay of HLA-A2-restricted CD8 + T cell responses against 11 selected peptides in PBMCs from 26 patients with mild, 8 with moderate, and 8 with severe COVID-19, respectively. (B) Log10-transformed mean spot-forming cell (SFC) counts (log10[Mean SFC +1]) per patient across 11 peptides in patients with mild, moderate, and severe COVID-19. Each point represents the mean response for one patient across 11 peptides ( n = 42 patients). A +1 pseudocount is added before the log10 transformation to accommodate zero values. Statistical comparisons were performed using a negative binomial mixed-effects model with Holm-Bonferroni correction for multiple comparisons. NS: not significant. (C) Longitudinal ELISpot assay of HLA-A2-restricted CD8 + T cell responses against selected peptides in PBMCs from patients with mild and moderate COVID-19 (as indicated).

Article Snippet: anti-human-IFN-γ antibody , MABTECH , Cat# 3420-3-250; RRID: AB_907283.

Techniques: Enzyme-linked Immunospot, Transformation Assay

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