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Jackson Laboratory ot1 tcr transgenic mice
Ot1 Tcr Transgenic Mice, supplied by Jackson Laboratory, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/tcr+transgenic+mice/us12648963-606-6-20?v=Jackson+Laboratory
Average 86 stars, based on 1 article reviews

ot1 tcr transgenic mice - by Bioz Stars, 2026-08

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CD100 deficiency impairs proliferation, activation, and effector function of HBcAg-specific CD8 + T cells upon antigen stimulation in vitro . (A) Experimental scheme: Splenocytes from WT C93-TCR-transgenic and CD100KO C93-TCR-transgenic mice were stimulated with Cor93 peptide for 96 hours. (B) The absolute number of CD8 + T cells at each time point are shown. (C–F) Expression levels of Ki67, CD25, Eomes and T-bet in CD8 + T cells were analyzed by flow cytometry. (G) Frequency and mean fluorescence intensity (MFI) of IFN-γ in CD8 + T cells are displayed. Data are depicted as arithmetic means ± SEM. Differences between two groups were analyzed using unpaired Student’s t tests. KO, knockout.

Journal: Frontiers in Immunology

Article Title: Cell-autonomous co-stimulatory function of membrane-bound CD100 promotes activation and differentiation of HBcAg-specific CD8 + T cells

doi: 10.3389/fimmu.2026.1826036

Figure Lengend Snippet: CD100 deficiency impairs proliferation, activation, and effector function of HBcAg-specific CD8 + T cells upon antigen stimulation in vitro . (A) Experimental scheme: Splenocytes from WT C93-TCR-transgenic and CD100KO C93-TCR-transgenic mice were stimulated with Cor93 peptide for 96 hours. (B) The absolute number of CD8 + T cells at each time point are shown. (C–F) Expression levels of Ki67, CD25, Eomes and T-bet in CD8 + T cells were analyzed by flow cytometry. (G) Frequency and mean fluorescence intensity (MFI) of IFN-γ in CD8 + T cells are displayed. Data are depicted as arithmetic means ± SEM. Differences between two groups were analyzed using unpaired Student’s t tests. KO, knockout.

Article Snippet: HBV Cor93 TCR-transgenic (C93-TCRtg) mice were purchased from the Jackson Laboratory.

Techniques: Activation Assay, In Vitro, Transgenic Assay, Expressing, Flow Cytometry, Fluorescence, Knock-Out

mCD100 deficiency results in decreased proliferation and effector function of HBcAg-specific CD8 + T cells upon antigen stimulation in vitro . (A) Experimental setup: CD8 + T cells were isolated from WT C93-TCR-transgenic and CD100KO C93-TCR-transgenic mice, mixed at a 1:1 ratio, and cocultured with WT non-CD8 splenocytes in the presence of Cor93 peptide for 96 hours. (B) Frequency of CD8 + T cells. (C–E) Ki67, Eomes and T-bet expression on CD8 + T cells were detected by flow cytometry. (F–G) Frequencies and mean fluorescence intensity (MFI) of IFN-γ and Granzyme B in CD8 + T cells are shown. Data are depicted as arithmetic means ± SEM. Differences between two groups were analyzed using unpaired Student’s t tests. MACS, Magnetically Activated Cell Sorting; KO, knockout.

Journal: Frontiers in Immunology

Article Title: Cell-autonomous co-stimulatory function of membrane-bound CD100 promotes activation and differentiation of HBcAg-specific CD8 + T cells

doi: 10.3389/fimmu.2026.1826036

Figure Lengend Snippet: mCD100 deficiency results in decreased proliferation and effector function of HBcAg-specific CD8 + T cells upon antigen stimulation in vitro . (A) Experimental setup: CD8 + T cells were isolated from WT C93-TCR-transgenic and CD100KO C93-TCR-transgenic mice, mixed at a 1:1 ratio, and cocultured with WT non-CD8 splenocytes in the presence of Cor93 peptide for 96 hours. (B) Frequency of CD8 + T cells. (C–E) Ki67, Eomes and T-bet expression on CD8 + T cells were detected by flow cytometry. (F–G) Frequencies and mean fluorescence intensity (MFI) of IFN-γ and Granzyme B in CD8 + T cells are shown. Data are depicted as arithmetic means ± SEM. Differences between two groups were analyzed using unpaired Student’s t tests. MACS, Magnetically Activated Cell Sorting; KO, knockout.

Article Snippet: HBV Cor93 TCR-transgenic (C93-TCRtg) mice were purchased from the Jackson Laboratory.

Techniques: In Vitro, Isolation, Transgenic Assay, Expressing, Flow Cytometry, Fluorescence, FACS, Knock-Out

Characterization of transcriptomic alterations in CD100KO HBcAg-specific CD8 + T cells. (A) Experimental setup: Splenocytes from WT and CD100KO C93-TCR-transgenic mice were stimulated with Cor93 peptide for 48 hours, followed by CD8 + T cell isolation for RNA sequencing. (B) The number of DEGs between the two groups. (C) Top 20 KEGG enriched terms for significant regulated genes between two groups. (D) Heatmap depicting the expression levels of significant DEGs involved in the “cytokine-cytokine receptor interaction” KEGG pathway. (E) GO biological process terms for the top 20 hub genes (with highest connectivity) from the PPI network of DEGs in the “cytokine-cytokine receptor interaction” pathway. (F) GSEA reveals significant downregulation of gene sets associated with the PI3K-Akt, mTOR, NF-κB, and JAK-STAT signaling pathways in CD100KO CD8 + T cells. DEGs, differentially expressed genes; GSEA, gene set enrichment analysis; PPI, protein-protein interaction; FDR, false discovery rate; NES, normalized enrichment score.

Journal: Frontiers in Immunology

Article Title: Cell-autonomous co-stimulatory function of membrane-bound CD100 promotes activation and differentiation of HBcAg-specific CD8 + T cells

doi: 10.3389/fimmu.2026.1826036

Figure Lengend Snippet: Characterization of transcriptomic alterations in CD100KO HBcAg-specific CD8 + T cells. (A) Experimental setup: Splenocytes from WT and CD100KO C93-TCR-transgenic mice were stimulated with Cor93 peptide for 48 hours, followed by CD8 + T cell isolation for RNA sequencing. (B) The number of DEGs between the two groups. (C) Top 20 KEGG enriched terms for significant regulated genes between two groups. (D) Heatmap depicting the expression levels of significant DEGs involved in the “cytokine-cytokine receptor interaction” KEGG pathway. (E) GO biological process terms for the top 20 hub genes (with highest connectivity) from the PPI network of DEGs in the “cytokine-cytokine receptor interaction” pathway. (F) GSEA reveals significant downregulation of gene sets associated with the PI3K-Akt, mTOR, NF-κB, and JAK-STAT signaling pathways in CD100KO CD8 + T cells. DEGs, differentially expressed genes; GSEA, gene set enrichment analysis; PPI, protein-protein interaction; FDR, false discovery rate; NES, normalized enrichment score.

Article Snippet: HBV Cor93 TCR-transgenic (C93-TCRtg) mice were purchased from the Jackson Laboratory.

Techniques: Transgenic Assay, Cell Isolation, RNA Sequencing, Expressing, Protein-Protein interactions

( A-B ) Mouse CD8+ T cells were treated with indicated cytokines for 24 hours prior to RNA isolation for bulk RNAseq analysis; (A) Geneset enrichment analysis (GSEA) for DEGs between IL-2/21-Trikine-treated vs sIL2-treated mouse CD8+ T cells (left) and heatmap of leading-edge genes (right). The DEG ranks were projected onto two gene sets defined based on a previously published ChIP-seq data . pSTAT3-promoted genes, genes with stronger ChIP peaks in STAT3-WT mice. (B) Relative expression of stemness genes Tcf7 , Bach2 , Il7r , Slamf6 , and Sell between mouse CD8+ T cells treated with either sIL-2 or IL-2/21-Trikine. Results from one independent experiment with 3 technical replicates. ( C-F ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8+ T cells (1 x 10 7 ) that were preconditioned for 7 days with sIL-2, sIL-2 + IL-21, or IL-2/21-Trikine on day 5, followed by i.p. injections of cytokine (5 μg functional cytokine) every other day until day 12 (n = 5 animals); the experimental timeline (C), average tumor growth curves (D), average weight change curves (E), and survival curves (F). Results from one independent experiment. ( G-I ) Healthy C57BL/6 mice were dosed with 10 μg functional dose of sIL-2, IL-21, sIL-2 + IL-21, IL-2/21-Trikine or PBS every other day for four total doses (n = 10 mice); experimental timeline (G) percent weight change at study endpoint (H), serum amyloid A concentrations at study endpoint (I). Results are combined from 2 independent experiments. ( J-M ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8+ T cells (1 x 10 7 ) that were preconditioned for 7 days with sIL-2, sIL-2 + IL-21, or IL-2/21-Trikine on day 5, followed by i.p. injections of cytokine (2.5 μg functional cytokine) every other day until day 12 (n = 14/15 animals); the experimental timeline (J), average tumor growth curves (K), average weight change curves (L), and survival curves (M). Results are combined from 2 independent experiments. ( N ) Proportions of pmel T cell memory populations, TCM and TEM in the draining lymph node 16 days post tumor injection from mice receiving low dose regimen (J-M) (n = 8). Results representative of 2 independent experiments. ( O-P ) MFI of Sca1 (O) and IL7Ra (P) cells in the draining lymph node 12 days post ACT compared to on the day of ACT from mice receiving low dose regimen (J) (n = 8). Results representative of 2 independent experiments. ( Q ) Intratumoral, live Thy1.1+CD8+ cells were sorted from dissociated tumor 12 days after ACT as in (J) and sent for scRNA-seq. 8 transcriptionally distinct clusters are seen by UMAP (left) and cluster composition by treatment is displayed (right). scRNAseq data is from one independent experiment. Schematics in C, G, and J created using BioRender.com .

Journal: Science (New York, N.Y.)

Article Title: Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

doi: 10.1126/science.adx9954

Figure Lengend Snippet: ( A-B ) Mouse CD8+ T cells were treated with indicated cytokines for 24 hours prior to RNA isolation for bulk RNAseq analysis; (A) Geneset enrichment analysis (GSEA) for DEGs between IL-2/21-Trikine-treated vs sIL2-treated mouse CD8+ T cells (left) and heatmap of leading-edge genes (right). The DEG ranks were projected onto two gene sets defined based on a previously published ChIP-seq data . pSTAT3-promoted genes, genes with stronger ChIP peaks in STAT3-WT mice. (B) Relative expression of stemness genes Tcf7 , Bach2 , Il7r , Slamf6 , and Sell between mouse CD8+ T cells treated with either sIL-2 or IL-2/21-Trikine. Results from one independent experiment with 3 technical replicates. ( C-F ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8+ T cells (1 x 10 7 ) that were preconditioned for 7 days with sIL-2, sIL-2 + IL-21, or IL-2/21-Trikine on day 5, followed by i.p. injections of cytokine (5 μg functional cytokine) every other day until day 12 (n = 5 animals); the experimental timeline (C), average tumor growth curves (D), average weight change curves (E), and survival curves (F). Results from one independent experiment. ( G-I ) Healthy C57BL/6 mice were dosed with 10 μg functional dose of sIL-2, IL-21, sIL-2 + IL-21, IL-2/21-Trikine or PBS every other day for four total doses (n = 10 mice); experimental timeline (G) percent weight change at study endpoint (H), serum amyloid A concentrations at study endpoint (I). Results are combined from 2 independent experiments. ( J-M ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8+ T cells (1 x 10 7 ) that were preconditioned for 7 days with sIL-2, sIL-2 + IL-21, or IL-2/21-Trikine on day 5, followed by i.p. injections of cytokine (2.5 μg functional cytokine) every other day until day 12 (n = 14/15 animals); the experimental timeline (J), average tumor growth curves (K), average weight change curves (L), and survival curves (M). Results are combined from 2 independent experiments. ( N ) Proportions of pmel T cell memory populations, TCM and TEM in the draining lymph node 16 days post tumor injection from mice receiving low dose regimen (J-M) (n = 8). Results representative of 2 independent experiments. ( O-P ) MFI of Sca1 (O) and IL7Ra (P) cells in the draining lymph node 12 days post ACT compared to on the day of ACT from mice receiving low dose regimen (J) (n = 8). Results representative of 2 independent experiments. ( Q ) Intratumoral, live Thy1.1+CD8+ cells were sorted from dissociated tumor 12 days after ACT as in (J) and sent for scRNA-seq. 8 transcriptionally distinct clusters are seen by UMAP (left) and cluster composition by treatment is displayed (right). scRNAseq data is from one independent experiment. Schematics in C, G, and J created using BioRender.com .

Article Snippet: TCR-transgenic Thy1.1 + pmel-1 (pmel) mice (B6.Cg-Thy1a/Cy Tg(TcraTcrb)8Rest/J) were originally purchased from the Jackson Laboratory and maintained in the Stanford University-Lorry Lokey (SIM1) Facility.

Techniques: Isolation, RNA sequencing, ChIP-sequencing, Expressing, Functional Assay, Injection

( A-C ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8 + T cells (5 × 10 6 ) on day 5, followed by i.p. injections of cytokine (3 μg functional cytokine) or PBS every other day until day 20 (n = 5 animals); the experimental timeline (A), average tumor growth curves (B), and survival curves (C). Results representative of 2 independent experiments. ( D-I ) Experimental setting is described in . C57BL/6 mice bearing established s.c. B16F10 tumors received i.v. ACT of pmel CD8 + T cells (5 × 10 6 ) on day 9, followed by i.p. injections of cytokine (3 μg functional cytokine) or left NT on days 9 and 11 (n = 5 animals). Mice were sacrificed on day 13, tumors were collected for flow cytometry analysis. Frequencies of Thy1.1 + CD8 + tumor-infiltrating lymphocytes (TILs) among single-live cells (D), frequencies of SCA-1 + CD62L + cells among Thy1.1 + CD8 + TILs (E), frequencies of PD-1+CD44+ cells among Thy1.1+CD8+ TILs (F) frequencies of Granzyme B+ cells among PD-1+CD44+ Thy1.1+CD8+ TILs (G), frequencies of IFN-γ+ cells among PD-1+CD44+ Thy1.1+CD8+ TILs (H), frequencies of TCF1+TIM-3- cells among PD-1+CD44+ Thy1.1+CD8+ TILs (I). Results representative of 2 independent experiments. ( J-L ), C57BL/6 mice bearing established s.c. B16F10 tumors (n = 5-10 animals) were administered cytokine (3 μg functional cytokine), PBS, or left NT i.p. on day 5 and every other day until day 19. For combination therapy, anti-PD-1 antibody (200 μg per dose) were i.p. administered on days 7, 11, 15, and 19; the experimental timeline (J), average tumor growth curves (K), survival curves (L). Results representative of 2 independent experiments. ( M ) Schematic of patient-derived organoid experiment. ( N ) Percentages of T133 tumor organoid viability in experiment as in (M). Results representative of 2 independent experiments. All data represent mean ± s.e.m. and are analyzed by one-way (D-I, N) or two-way (B, K) ANOVA with Tukey’s post-test. Schematics in D and M created using BioRender.com .

Journal: Science (New York, N.Y.)

Article Title: Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

doi: 10.1126/science.adx9954

Figure Lengend Snippet: ( A-C ), C57BL/6 mice bearing established s.c. B16F10 tumors received intravenous (i.v.) adoptive cell transfer (ACT) of pmel CD8 + T cells (5 × 10 6 ) on day 5, followed by i.p. injections of cytokine (3 μg functional cytokine) or PBS every other day until day 20 (n = 5 animals); the experimental timeline (A), average tumor growth curves (B), and survival curves (C). Results representative of 2 independent experiments. ( D-I ) Experimental setting is described in . C57BL/6 mice bearing established s.c. B16F10 tumors received i.v. ACT of pmel CD8 + T cells (5 × 10 6 ) on day 9, followed by i.p. injections of cytokine (3 μg functional cytokine) or left NT on days 9 and 11 (n = 5 animals). Mice were sacrificed on day 13, tumors were collected for flow cytometry analysis. Frequencies of Thy1.1 + CD8 + tumor-infiltrating lymphocytes (TILs) among single-live cells (D), frequencies of SCA-1 + CD62L + cells among Thy1.1 + CD8 + TILs (E), frequencies of PD-1+CD44+ cells among Thy1.1+CD8+ TILs (F) frequencies of Granzyme B+ cells among PD-1+CD44+ Thy1.1+CD8+ TILs (G), frequencies of IFN-γ+ cells among PD-1+CD44+ Thy1.1+CD8+ TILs (H), frequencies of TCF1+TIM-3- cells among PD-1+CD44+ Thy1.1+CD8+ TILs (I). Results representative of 2 independent experiments. ( J-L ), C57BL/6 mice bearing established s.c. B16F10 tumors (n = 5-10 animals) were administered cytokine (3 μg functional cytokine), PBS, or left NT i.p. on day 5 and every other day until day 19. For combination therapy, anti-PD-1 antibody (200 μg per dose) were i.p. administered on days 7, 11, 15, and 19; the experimental timeline (J), average tumor growth curves (K), survival curves (L). Results representative of 2 independent experiments. ( M ) Schematic of patient-derived organoid experiment. ( N ) Percentages of T133 tumor organoid viability in experiment as in (M). Results representative of 2 independent experiments. All data represent mean ± s.e.m. and are analyzed by one-way (D-I, N) or two-way (B, K) ANOVA with Tukey’s post-test. Schematics in D and M created using BioRender.com .

Techniques: Functional Assay, Flow Cytometry, Derivative Assay

A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into LM TAG -infected C57BL/6 (B6; Thy1.2) or tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. TCR TAG were re-isolated from spleens and livers of infected mice (green) and tumor mice (blue) for flow cytometric analysis between 2.5-70+ days post-transfer. B. TCR TAG cell numbers in the spleens (left) and livers (right) of LM TAG -infected mice and tumor-bearing mice. Dots represent mean and error bars represent SEM. For B-D and F n = 2-3 mice per group (infection); n = 3 mice per group (tumor). Data is representative of two independent experiments. C. Histograms (left) showing TCR TAG PD1 expression from infection shown in comparison to naive (N; gray). For C-D and F , each symbol represents an individual mouse, and one-sample Student’s t -test with Bonferroni correction was used to determine significance: *, P <0.008 (infection); *, P <0.007 (tumor); ns, not statistically significant. D. Histograms and summary plots of CD38 expression and CD101 expression in spleens and livers from tumor mice shown in comparison to N. E. TCR TAG IFNγ and TNFα production after 4-hour ex vivo TAG peptide stimulation, with inset numbers indicating the percentage of cells in each gate. Gates were set based on no peptide stimulation controls. Right, summary plots of % IFNγ + TCR TAG after peptide stimulation; each symbol represents an individual mouse; n = 3-4 mice per group. *, P <0.05; **, P <0.01; ****, P <0.0001 (one-way ANOVA with post hoc Tukey test). F. Histograms and summary plots of CD122 expression and BCL2 expression from tumor mice shown in comparison to N. All flow plots are gated on live CD8 + Thy1.1 + cells, and flow data for each timepoint is concatenated from all biological replicates. Summary plots (right) show MFI (geometric m ean fluorescence intensity).

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into LM TAG -infected C57BL/6 (B6; Thy1.2) or tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. TCR TAG were re-isolated from spleens and livers of infected mice (green) and tumor mice (blue) for flow cytometric analysis between 2.5-70+ days post-transfer. B. TCR TAG cell numbers in the spleens (left) and livers (right) of LM TAG -infected mice and tumor-bearing mice. Dots represent mean and error bars represent SEM. For B-D and F n = 2-3 mice per group (infection); n = 3 mice per group (tumor). Data is representative of two independent experiments. C. Histograms (left) showing TCR TAG PD1 expression from infection shown in comparison to naive (N; gray). For C-D and F , each symbol represents an individual mouse, and one-sample Student’s t -test with Bonferroni correction was used to determine significance: *, P <0.008 (infection); *, P <0.007 (tumor); ns, not statistically significant. D. Histograms and summary plots of CD38 expression and CD101 expression in spleens and livers from tumor mice shown in comparison to N. E. TCR TAG IFNγ and TNFα production after 4-hour ex vivo TAG peptide stimulation, with inset numbers indicating the percentage of cells in each gate. Gates were set based on no peptide stimulation controls. Right, summary plots of % IFNγ + TCR TAG after peptide stimulation; each symbol represents an individual mouse; n = 3-4 mice per group. *, P <0.05; **, P <0.01; ****, P <0.0001 (one-way ANOVA with post hoc Tukey test). F. Histograms and summary plots of CD122 expression and BCL2 expression from tumor mice shown in comparison to N. All flow plots are gated on live CD8 + Thy1.1 + cells, and flow data for each timepoint is concatenated from all biological replicates. Summary plots (right) show MFI (geometric m ean fluorescence intensity).

Article Snippet: TCR TAG transgenic mice (B6.Cg-Tg(TcraY1,TcrbY1)416Tev/J, RRID:IMSR_JAX:005236) , Cre-ER T2 mice (B6.129-Gt(ROSA)26Sortm1(cre/ERT2)Tyj/J, RRID:IMSR_JAX:008463), Rosa26-Cas9 mice (Gt(ROSA)26Sortm1.1(CAG-cas9*,-EGFP)Fezh/J, RRID:IMSR_JAX:026179), B6 Thy1.1 mice (B6.PL-Thy1a/CyJ, RRID:IMSR_JAX:000406), and B6 mice (C57BL/6J, RRID:IMSR_JAX:000664) were purchased from the Jackson Laboratory.

Techniques: Infection, Isolation, Expressing, Comparison, Ex Vivo, Fluorescence

A. Experimental setup as shown in . Histograms and summary plots of CD44 expression from infection (green) and tumor (blue) in spleen and liver shown in comparison to naive (N; gray). Each symbol represents an individual mouse, and one-sample Student’s t -test with Bonferroni correction was used to determine significance: *, P <0.008 (infection); *, P <0.007 (tumor); ns, not statistically significant. n = 2-3 mice per group (infection); n = 3 mice per group (tumor). Data is representative of two independent experiments. B. TCR TAG IFNγ and TNFα production after 4-hour ex vivo TAG peptide stimulation, with inset numbers indicating the percentage of cells in each gate. Gates were set based on no peptide stimulation controls. Summary plots show % IFNγ + TCR TAG after peptide stimulation; each symbol represents an individual mouse; n = 3-4 mice per group. ****, P <0.0001 (two-way ANOVA followed by post hoc Šídák’s multiple comparisons test). All flow plots are gated on live CD8 + Thy1.1 + cells and flow data for each timepoint is concatenated from all biological replicates. Data is representative of two independent experiments.

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Experimental setup as shown in . Histograms and summary plots of CD44 expression from infection (green) and tumor (blue) in spleen and liver shown in comparison to naive (N; gray). Each symbol represents an individual mouse, and one-sample Student’s t -test with Bonferroni correction was used to determine significance: *, P <0.008 (infection); *, P <0.007 (tumor); ns, not statistically significant. n = 2-3 mice per group (infection); n = 3 mice per group (tumor). Data is representative of two independent experiments. B. TCR TAG IFNγ and TNFα production after 4-hour ex vivo TAG peptide stimulation, with inset numbers indicating the percentage of cells in each gate. Gates were set based on no peptide stimulation controls. Summary plots show % IFNγ + TCR TAG after peptide stimulation; each symbol represents an individual mouse; n = 3-4 mice per group. ****, P <0.0001 (two-way ANOVA followed by post hoc Šídák’s multiple comparisons test). All flow plots are gated on live CD8 + Thy1.1 + cells and flow data for each timepoint is concatenated from all biological replicates. Data is representative of two independent experiments.

Techniques: Expressing, Infection, Comparison, Ex Vivo

A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. TCR TAG were re-isolated from spleens and livers for flow cytometric analysis at 2.5, 5, 10, 14, 28, 35, and 70+ days post-transfer. B. Histograms and summary plots of KI67 expression of TCR TAG in spleens and livers from tumor-bearing mice shown in comparison to N, with % KI67 + gate set to exclude N. Flow plots for each time point were concatenated from all biological replicates; each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. C. Left, representative dot plots of KI67 expression and DNA content staining in TCR TAG in spleens and livers of AST;Cre-ER T2 mice, with inset numbers indicating the percentage of cells in G 0 , G 1 , and S-G 2 M phases of cell cycle. Middle, summary plots of the percentage of TCR TAG in G 0 , G 1 , and S-G 2 M, shown as mean ± SEM. Right, summary plots of the percentage of TCR TAG in S-G 2 M. Each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. D. Histograms and summary plots of TCF1 expression in spleens and livers, with TCF1 + gate shown. Flow plots for each time point were concatenated from all biological replicates; each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. E. Longitudinal analysis of the absolute cell numbers of TCR TAG cells (blue) in the spleens and livers of tumor-bearing mice and the % TCF1 + TCR TAG (yellow), shown as mean ± SEM. F. Summary plots of the % TCF1 + (filled bars) and TCF1 − (open bars) TCR TAG within the S-G 2 /M + subset, shown as mean ± SEM. Data is representative of two independent experiments with n = 3 mice per group

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. TCR TAG were re-isolated from spleens and livers for flow cytometric analysis at 2.5, 5, 10, 14, 28, 35, and 70+ days post-transfer. B. Histograms and summary plots of KI67 expression of TCR TAG in spleens and livers from tumor-bearing mice shown in comparison to N, with % KI67 + gate set to exclude N. Flow plots for each time point were concatenated from all biological replicates; each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. C. Left, representative dot plots of KI67 expression and DNA content staining in TCR TAG in spleens and livers of AST;Cre-ER T2 mice, with inset numbers indicating the percentage of cells in G 0 , G 1 , and S-G 2 M phases of cell cycle. Middle, summary plots of the percentage of TCR TAG in G 0 , G 1 , and S-G 2 M, shown as mean ± SEM. Right, summary plots of the percentage of TCR TAG in S-G 2 M. Each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. D. Histograms and summary plots of TCF1 expression in spleens and livers, with TCF1 + gate shown. Flow plots for each time point were concatenated from all biological replicates; each symbol represents an individual mouse. *, P <0.007 determined by one-sample Student’s t -test with Bonferroni correction. E. Longitudinal analysis of the absolute cell numbers of TCR TAG cells (blue) in the spleens and livers of tumor-bearing mice and the % TCF1 + TCR TAG (yellow), shown as mean ± SEM. F. Summary plots of the % TCF1 + (filled bars) and TCF1 − (open bars) TCR TAG within the S-G 2 /M + subset, shown as mean ± SEM. Data is representative of two independent experiments with n = 3 mice per group

Techniques: Isolation, Expressing, Comparison, Staining

A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into LM TAG -infected B6 (Thy1.2) or tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice and 5-ethynyl-2’-deoxyurdine (EdU) and 5-bromo-2’-deoxyuridine (BrdU) were administered on days 35 and 42, respectively. TCR TAG were re-isolated on day 43 from spleens of infected mice (green) and spleens and livers of tumor-bearing mice (blue) for flow cytometric analysis. B. Schematic representation of the expected distributions of EdU and BrdU incorporation for T cells proliferating in a progenitor-progeny hierarchy (left) or stochastically (right). C. Top, dot plots of TCR TAG EdU and BrdU incorporation in infected mice and tumor-bearing mice. Unlabeled cells shown in gray with inset numbers showing the percentage of nucleoside labeled cells in each gate. Bottom, summary plots of % EdU + (red), EdU + ,BrdU + (purple), and BrdU + (cyan) TCR TAG within each nucleoside labeled subset. Each symbol represents an individual mouse. **, P <0.01 determined by repeated measures, one-way ANOVA with post hoc Tukey test. D. Concatenated histogram of TCF1 expression in nucleoside-labeled TCR TAG and summary plot of the percentage of nucleoside-labeled TCR TAG expressing TCF1. Each symbol represents an individual mouse. *, P <0.025 determined by one-sample Student’s t -test with Bonferroni correction with n=4-5 mice per group and representative of two independent experiments. Flow plots show data concatenated from all biological replicates.

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into LM TAG -infected B6 (Thy1.2) or tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice and 5-ethynyl-2’-deoxyurdine (EdU) and 5-bromo-2’-deoxyuridine (BrdU) were administered on days 35 and 42, respectively. TCR TAG were re-isolated on day 43 from spleens of infected mice (green) and spleens and livers of tumor-bearing mice (blue) for flow cytometric analysis. B. Schematic representation of the expected distributions of EdU and BrdU incorporation for T cells proliferating in a progenitor-progeny hierarchy (left) or stochastically (right). C. Top, dot plots of TCR TAG EdU and BrdU incorporation in infected mice and tumor-bearing mice. Unlabeled cells shown in gray with inset numbers showing the percentage of nucleoside labeled cells in each gate. Bottom, summary plots of % EdU + (red), EdU + ,BrdU + (purple), and BrdU + (cyan) TCR TAG within each nucleoside labeled subset. Each symbol represents an individual mouse. **, P <0.01 determined by repeated measures, one-way ANOVA with post hoc Tukey test. D. Concatenated histogram of TCF1 expression in nucleoside-labeled TCR TAG and summary plot of the percentage of nucleoside-labeled TCR TAG expressing TCF1. Each symbol represents an individual mouse. *, P <0.025 determined by one-sample Student’s t -test with Bonferroni correction with n=4-5 mice per group and representative of two independent experiments. Flow plots show data concatenated from all biological replicates.

Techniques: Infection, Isolation, BrdU Incorporation Assay, Labeling, Expressing

A. Histogram of TCF1 expression 48 hours post transfection of naive Cas9;TCR TAG with sgRNA transfected with non-targeting control sgRNA (NTC; light green) or TCF7 sgRNA (TCF1KO; dark teal), data representative of two independent experiments. B. Experimental scheme: NTC or TCF1KO TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. EdU was administered in the final 3 days. TCR TAG were re-isolated on day 35 from the spleens and livers for flow cytometric analysis. C. NTC and TCF1KO TCR TAG cell numbers recovered from tumor-bearing mice. D. Representative dot plots of KI67 expression and EdU incorporation of NTC and TCF1KO TCR TAG cells, with gates set based on KI67 − /EdU − cells. E. Top, summary plots of the percentage (left) and absolute number (right) of EdU + NTC or TCF1KO TCR TAG . Bottom, summary plots of the percentage (left) and absolute number (right) of KI67 + or TCF1KO TCR TAG . For C and E , each symbol represents an individual mouse, with two-way ANOVA followed by post hoc Šídák’s multiple comparisons test with n=4-5 mice per group and representative of two independent experiments.

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Histogram of TCF1 expression 48 hours post transfection of naive Cas9;TCR TAG with sgRNA transfected with non-targeting control sgRNA (NTC; light green) or TCF7 sgRNA (TCF1KO; dark teal), data representative of two independent experiments. B. Experimental scheme: NTC or TCF1KO TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. EdU was administered in the final 3 days. TCR TAG were re-isolated on day 35 from the spleens and livers for flow cytometric analysis. C. NTC and TCF1KO TCR TAG cell numbers recovered from tumor-bearing mice. D. Representative dot plots of KI67 expression and EdU incorporation of NTC and TCF1KO TCR TAG cells, with gates set based on KI67 − /EdU − cells. E. Top, summary plots of the percentage (left) and absolute number (right) of EdU + NTC or TCF1KO TCR TAG . Bottom, summary plots of the percentage (left) and absolute number (right) of KI67 + or TCF1KO TCR TAG . For C and E , each symbol represents an individual mouse, with two-way ANOVA followed by post hoc Šídák’s multiple comparisons test with n=4-5 mice per group and representative of two independent experiments.

Techniques: Expressing, Transfection, Control, Isolation

A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. FTY720 treatment or vehicle was administered every other day beginning on day −1 for 6 days or day 28 for 3 weeks. EdU was administered for the final 3 days prior to harvest. TCR TAG were re-isolated on day 5 and day 49 from the livers for flow cytometric analysis. B. Representative histogram and summary plot of % CD3ε + cells in peripheral blood 24 hours after initiation of FTY720 (purple) or vehicle (black) treatment. ****, P <000.1 determined by unpaired, two-tailed Student’s t -test. C. TCR TAG cell numbers in the livers of vehicle- (open black bars) and FTY720 (purple bars)-treated mice. Each symbol represents an individual mouse, with ****, P <0.0001 determined by two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. D. Representative histogram of EdU+ incorporation in TCR TAG T cells and summary plot of % EdU + from the livers of vehicle- and FTY720-treated AST;Cre-ER T2 mice. Each symbol represents an individual mouse, two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. E. Left, representative dot plots of TCF1 and KI67 expression in EdU + TCR TAG from the livers of AST;CreER T2 mice at day 5 (top) and day 49 (bottom) following vehicle and FTY720 treatment. Right, summary plots of % TCF1 hi (top) and KI67 + (bottom) of EdU + TCR TAG . Each symbol represents an individual mouse, two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. For B-E , n=4-5 mice per group and representative of two independent experiments.

Journal: bioRxiv

Article Title: TCF1 lo CD8 T cells proliferate and persist autonomously in tumors

doi: 10.64898/2026.01.17.700120

Figure Lengend Snippet: A. Experimental scheme: naive TCR TAG (Thy1.1) were adoptively transferred into tamoxifen-treated AST;Cre-ER T2 (Thy1.2) mice. FTY720 treatment or vehicle was administered every other day beginning on day −1 for 6 days or day 28 for 3 weeks. EdU was administered for the final 3 days prior to harvest. TCR TAG were re-isolated on day 5 and day 49 from the livers for flow cytometric analysis. B. Representative histogram and summary plot of % CD3ε + cells in peripheral blood 24 hours after initiation of FTY720 (purple) or vehicle (black) treatment. ****, P <000.1 determined by unpaired, two-tailed Student’s t -test. C. TCR TAG cell numbers in the livers of vehicle- (open black bars) and FTY720 (purple bars)-treated mice. Each symbol represents an individual mouse, with ****, P <0.0001 determined by two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. D. Representative histogram of EdU+ incorporation in TCR TAG T cells and summary plot of % EdU + from the livers of vehicle- and FTY720-treated AST;Cre-ER T2 mice. Each symbol represents an individual mouse, two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. E. Left, representative dot plots of TCF1 and KI67 expression in EdU + TCR TAG from the livers of AST;CreER T2 mice at day 5 (top) and day 49 (bottom) following vehicle and FTY720 treatment. Right, summary plots of % TCF1 hi (top) and KI67 + (bottom) of EdU + TCR TAG . Each symbol represents an individual mouse, two-way ANOVA followed by post hoc Šídák’s multiple comparisons test. For B-E , n=4-5 mice per group and representative of two independent experiments.

Techniques: Isolation, Two Tailed Test, Expressing

(A, B) WT, heterozygous (±), and homozygous CD8beta-LSSmOrange (++) mice were intranasally infected with influenza, and on day 10, cells were isolated from lungs from uninfected (Un, open symbols) and infected (Inf, filled symbols) mice and analyzed by flow cytometry. (A, B) Number of lung tissue–resident CD8 T cells (left) and CD4 T cells (right) is shown in (A), and the percentage of lung-resident CD8 T cells expressing activation markers in influenza-infected mice is shown in (B). Data are combined from two separate experiments, n = 2–8. There were no significant differences between WT, +/−, or ++ infected mice. (C, D) T cells were isolated from heterozygous CD8beta-LSSmOrange OTI TCR transgenic and from CAG-GFP OTI TCR transgenic mice, and co-adoptively transferred into WT mice, and the recipient mice were infected with recombinant influenza expressing OVA peptide and analyzed by flow cytometry 10 d after infection. (C, D) Ratio of WT CAG-GFP OTI to CD8-LSSmOrange OTI T cells in spleen, draining lymph nodes, and lung tissue is shown in (C), and the expression of activation markers on lung-resident WT OTI T cells (GFP), CD8beta-LSSmOrange OTI T cells (LSSm), and endogenous (non-OT1) CD8 cells (Endog) is shown in (D). (C) Data are from three individual mice; the ratio in spleen, but not lymph node or lung, is significantly different from the starting ratio of 1.02 (* P < 0.05) in (C); and the expression of activation markers in the WT and CD8-LSSm-Orange OT1 cells is not significantly different.

Journal: Life Science Alliance

Article Title: Intravital imaging of the formation and resolution of MHC class II–positive T-cell activation niches

doi: 10.26508/lsa.202503476

Figure Lengend Snippet: (A, B) WT, heterozygous (±), and homozygous CD8beta-LSSmOrange (++) mice were intranasally infected with influenza, and on day 10, cells were isolated from lungs from uninfected (Un, open symbols) and infected (Inf, filled symbols) mice and analyzed by flow cytometry. (A, B) Number of lung tissue–resident CD8 T cells (left) and CD4 T cells (right) is shown in (A), and the percentage of lung-resident CD8 T cells expressing activation markers in influenza-infected mice is shown in (B). Data are combined from two separate experiments, n = 2–8. There were no significant differences between WT, +/−, or ++ infected mice. (C, D) T cells were isolated from heterozygous CD8beta-LSSmOrange OTI TCR transgenic and from CAG-GFP OTI TCR transgenic mice, and co-adoptively transferred into WT mice, and the recipient mice were infected with recombinant influenza expressing OVA peptide and analyzed by flow cytometry 10 d after infection. (C, D) Ratio of WT CAG-GFP OTI to CD8-LSSmOrange OTI T cells in spleen, draining lymph nodes, and lung tissue is shown in (C), and the expression of activation markers on lung-resident WT OTI T cells (GFP), CD8beta-LSSmOrange OTI T cells (LSSm), and endogenous (non-OT1) CD8 cells (Endog) is shown in (D). (C) Data are from three individual mice; the ratio in spleen, but not lymph node or lung, is significantly different from the starting ratio of 1.02 (* P < 0.05) in (C); and the expression of activation markers in the WT and CD8-LSSm-Orange OT1 cells is not significantly different.

Article Snippet: For adoptive transfer experiments, OTII TCR transgenic mice (Jackson Laboratory) were crossed to CAG-mOFP mice ( ) and 1–4 × 10 6 CD4 Th1 cells were adoptively transferred into IEbeta-mAmetrine/CD11c-Venus/REX3 mice before immunization.

Techniques: Infection, Isolation, Flow Cytometry, Expressing, Activation Assay, Transgenic Assay, Recombinant

IEbeta-mAmetrine mice were crossed to CD11c-Venus and REX3 mice and were adoptively transferred with 1 × 10 6 in vitro–activated OTII Th1 cells expressing OFP. The mice were intradermally immunized with ovalbumin in CFA in the ear pinna, and the same area of tissue was sequentially imaged by two-photon microscopy on days 3, 6, 9, 13, 16, 19, and 23. (A) Relative position of hair follicles (dark circles) in the second harmonic generation image confirms imaging of the same location. (B) Clusters of CXCL10-BFP–expressing cells were identified by nearest neighbor analysis (20 cells within radius of 50 mm) and highlighted with blue border. (B, C) All class II–positive cells are shown in red, OTII Th1 cells are shown in white, and the outline of the cluster defined in (B) is shown with a blue outline. (C, D) Enlarged images of the data are shown in (C). Representative data from one of three experiments. (A, B, C, D) Bars are 100 μm in (A, B, C) and 20 μm (D). (E, F, G, H, I) CXCL10 clusters, OTII T cells, and class II–positive cells were identified, and the T cells and the class II+ cells expressing different combinations of CD11c and CXCL10 within and outside the CXCL10 clusters were determined. (E) Percentage of image area contained within the CXCL10 clusters. (F) Number of class II+ cells within the entire image. (G) Percentage of class II+ cells within the clusters. Total number of class II+ cells (black); class II+ only (green); class II+ and CD11c+ (yellow); class II+ and CXCL10+ (blue); and class II, CXCL10+, and CD11c+ (red). (H) Number of OTII Th1 cells within the entire image. (I) Percentage of T cells within the clusters. Data are the mean of three separate experiments with individual data points for each time point shown. Note there are only two data points for day 3 and day 6 and single data point for days 29 and 36.

Journal: Life Science Alliance

Article Title: Intravital imaging of the formation and resolution of MHC class II–positive T-cell activation niches

doi: 10.26508/lsa.202503476

Figure Lengend Snippet: IEbeta-mAmetrine mice were crossed to CD11c-Venus and REX3 mice and were adoptively transferred with 1 × 10 6 in vitro–activated OTII Th1 cells expressing OFP. The mice were intradermally immunized with ovalbumin in CFA in the ear pinna, and the same area of tissue was sequentially imaged by two-photon microscopy on days 3, 6, 9, 13, 16, 19, and 23. (A) Relative position of hair follicles (dark circles) in the second harmonic generation image confirms imaging of the same location. (B) Clusters of CXCL10-BFP–expressing cells were identified by nearest neighbor analysis (20 cells within radius of 50 mm) and highlighted with blue border. (B, C) All class II–positive cells are shown in red, OTII Th1 cells are shown in white, and the outline of the cluster defined in (B) is shown with a blue outline. (C, D) Enlarged images of the data are shown in (C). Representative data from one of three experiments. (A, B, C, D) Bars are 100 μm in (A, B, C) and 20 μm (D). (E, F, G, H, I) CXCL10 clusters, OTII T cells, and class II–positive cells were identified, and the T cells and the class II+ cells expressing different combinations of CD11c and CXCL10 within and outside the CXCL10 clusters were determined. (E) Percentage of image area contained within the CXCL10 clusters. (F) Number of class II+ cells within the entire image. (G) Percentage of class II+ cells within the clusters. Total number of class II+ cells (black); class II+ only (green); class II+ and CD11c+ (yellow); class II+ and CXCL10+ (blue); and class II, CXCL10+, and CD11c+ (red). (H) Number of OTII Th1 cells within the entire image. (I) Percentage of T cells within the clusters. Data are the mean of three separate experiments with individual data points for each time point shown. Note there are only two data points for day 3 and day 6 and single data point for days 29 and 36.

Techniques: In Vitro, Expressing, Microscopy, Imaging

(A, B, C, D, E, F, G, H) Replicate experiments of quantified in using 4 × 10 6 OTII Th1 cells and imaged on days 3, 9, 12, 16, 19, and 23 (A, B, C, D) and 1 × 10 6 OTII Th1 cells and imaged on days 6, 9, 13, 16, 19, 23,29, and 36 (E, F, G, H). (A, E) Second harmonic generation images. (B, F) Clusters of CXCL10-BFP–expressing cells highlighted with blue border. (C, G) All class II–positive cells in red, OTII Th1 cells in white, and the outline of the CXCL10-BFP cluster in blue are shown. (C, D, G, H) Enlarged images of the data shown in (C, G), respectively. (D, H) Bars are 100 μm, except in (D, H) when they are 20 μm.

Journal: Life Science Alliance

Article Title: Intravital imaging of the formation and resolution of MHC class II–positive T-cell activation niches

doi: 10.26508/lsa.202503476

Figure Lengend Snippet: (A, B, C, D, E, F, G, H) Replicate experiments of quantified in using 4 × 10 6 OTII Th1 cells and imaged on days 3, 9, 12, 16, 19, and 23 (A, B, C, D) and 1 × 10 6 OTII Th1 cells and imaged on days 6, 9, 13, 16, 19, 23,29, and 36 (E, F, G, H). (A, E) Second harmonic generation images. (B, F) Clusters of CXCL10-BFP–expressing cells highlighted with blue border. (C, G) All class II–positive cells in red, OTII Th1 cells in white, and the outline of the CXCL10-BFP cluster in blue are shown. (C, D, G, H) Enlarged images of the data shown in (C, G), respectively. (D, H) Bars are 100 μm, except in (D, H) when they are 20 μm.

Techniques: Expressing

Journal: Life Science Alliance

Article Title: Intravital imaging of the formation and resolution of MHC class II–positive T-cell activation niches

doi: 10.26508/lsa.202503476

Article Snippet: CD8beta-LSSmOrange mice were crossed to OTI TCR transgenic mice (Jackson Laboratory), and thymus and spleen were stained with antibodies to CD8, TCR Valpha2, and TCR Vbeta5 to confirm that LSSmOrange-positive CD8 T cells expressing the OTI TCR were appropriately selected.

Techniques: Infection, Isolation, Flow Cytometry, Expressing, Activation Assay, Transgenic Assay, Recombinant

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