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wild type fvb n mice  (Jackson Laboratory)

 
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    Jackson Laboratory wild type fvb n mice
    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Wild Type Fvb N 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/wild-type+mice/fvb+mice+nj/pmc13196419-230-1-7
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    Images

    1) Product Images from "CIT tumor lines: A series of immunogenic murine cutaneous squamous cell carcinoma cell lines derived from chemical carcinogenesis"

    Article Title: CIT tumor lines: A series of immunogenic murine cutaneous squamous cell carcinoma cell lines derived from chemical carcinogenesis

    Journal: JID Innovations

    doi: 10.1016/j.xjidi.2026.100477

    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant versus wild-type sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Figure Legend Snippet: CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant versus wild-type sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.

    Techniques Used: Immunopeptidomics, Selection, Binding Assay, Mutagenesis, Sequencing, Gene Expression, Generated, Enzyme-linked Immunospot, Isolation, Cell Culture, Negative Control

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    Article Snippet: Wild- type (WT) mice and Adnpmut mice (carrying a 14- nucleotide deletion in the C- terminus, Strain # 033128, Jackson Lab) with C57BL/6J background were utilized.

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    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
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    Charles River Laboratories wild type c57bl 6j male mice
    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Wild Type C57bl 6j Male Mice, supplied by Charles River Laboratories, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    press c57bl 6jrj wild type wt mice  (Janvier Labs)
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    Janvier Labs press c57bl 6jrj wild type wt mice
    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Press C57bl 6jrj Wild Type Wt Mice, supplied by Janvier Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    male c57bl 6ncrl wild type mice  (Charles River Laboratories)
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    Charles River Laboratories male c57bl 6ncrl wild type mice
    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Male C57bl 6ncrl Wild Type Mice, supplied by Charles River Laboratories, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    wild type c57bl 6 b6 mice  (Jackson Laboratory)
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    Jackson Laboratory wild type c57bl 6 b6 mice
    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant <t>versus</t> <t>wild-type</t> sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.
    Wild Type C57bl 6 B6 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
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    CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant versus wild-type sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.

    Journal: JID Innovations

    Article Title: CIT tumor lines: A series of immunogenic murine cutaneous squamous cell carcinoma cell lines derived from chemical carcinogenesis

    doi: 10.1016/j.xjidi.2026.100477

    Figure Lengend Snippet: CIT lines express identifiable neoantigens whose cognate CD8+ T cells are responsive to immunotherapy. ( a ) Selection criteria for CIT6 mutations predicted to generate neoantigens. (b) Predicted peptide log affinity ratio (affinity ratio is defined as the ratio between predicted binding affinity of mutant versus wild-type sequence) and gene expression level of mutations predicted to be top CIT6 candidate neoantigens, shown for H2-Dq and H2-Kq alleles. Affinity predictions were generated using the NetH2Pan algorithm. (c) Schematic of IFNγ ELISpot assay. For screening of CD8+ T cell and CD4+ T cell responses, CD8+ or CD4+ T cells were isolated from the spleens of tumor-bearing mice and co-cultured with total splenocytes from a healthy tumor-naïve mouse. (d) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Brinp3 peptide CPAFLPCTV (top row) and negative control (no peptide, bottom row). (e) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Brinp3 peptide CPAFLPCTV. Each colored line represents data points from a different mouse (n = 4 mice). (f) Representative IFNγ ELISpot images of the assay performed with CD8+ T cells from the spleen of CIT6-bearing mice cultured with 160ug/ml of Ttll4 peptide VPPSSLLPL (top row) and negative control (no peptide, bottom row). (g) Quantification of average IFNγ ELISpot spot counts for CD8+ T cells from the spleen of CIT6-bearing mice cultured with different concentrations of Ttll4 peptide VPPSSLLPL. Each colored line represents data points from a different mouse (n = 4 mice). (h) Spot counts from ELISpot assays performed with CD4+ T cells isolated from the spleens of CIT6-bearing mice and cultured with splenocytes from tumor-naïve mice loaded with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 4 mice). (i) Spot counts from ELISpot assays performed with total splenocytes from CIT6-bearing mice cultured with increasing concentrations of Brinp3 and Ttll4 peptides. Each colored line represents data points from a different mouse (n = 10 – 11 mice). (j) Schematic of ICI treatment regimen and ELISpot analysis. (k) Average spot counts of ICI-treated splenocytes cultured with 80 ug/ml of Ttll4 peptide vs negative control. Each line represents data points from a different mouse (n = 6 mice). Statistical analysis in panels e, g, h, i, and k was done by a paired t- test. ∗ P < .05, ∗∗ P < .01. Error bars in panels e, g, h and i represent mean and SD. CIT, carcinogen-induced tumor; ICI, Immune checkpoint inhibitor.

    Article Snippet: Eight-week-old wild-type FVB/N mice were purchased from Jackson Laboratories.

    Techniques: Immunopeptidomics, Selection, Binding Assay, Mutagenesis, Sequencing, Gene Expression, Generated, Enzyme-linked Immunospot, Isolation, Cell Culture, Negative Control