rabbit anti mouse primary cd8  (Cell Signaling Technology Inc)

Journal: Pharmaceuticals

Article Title: AHR Agonist ITE Boosted PD1 Antibody’s Effects by Inhibiting Myeloid-Derived Cells Suppressive Cells in an Orthotopic Mouse Glioma Model

doi: 10.3390/ph18040471

Figure Lengend Snippet: Synergistic effect of ITE and PD1 antibody on cytotoxic immune cell infiltration. ( A,B ) Gating Strategy for myeloid cells. R2 represents single cells, R3 represents live cells, R4 represents CD45+ immune cells, R5 represents non-neutrophils, and R6 represents myeloid cells. R7 was composed of CD11c+ lymphocytes, NK cells, and monocytes. ( C – G ) Effects of DMSO, ITE, PD1 antibody, and ITE+PD1 antibody treatments on immune cells in brain tumors and spleen. ( C ) Percentage of CD45+ cells in brain tumors and spleen (* p < 0.05). ( D ) Percentage of CD4-CD8+T cells in brain tumors and spleen (* p < 0.05). ( E ) Immunofluorescence and immunohistochemical staining of frozen brain tumor sections to detect CD8 T-cell antigens; blue: cell nucleus, red: CD8. ( F ) Detection of IFN-γ in brain by ELISA. ( G ) Percentage of CD4+T cells in brain tumors and spleen (** p < 0.05). ( H ) Immunofluorescence and immunohistochemical staining of frozen brain tumor sections to detect CD4 T-cell antigens; blue: cell nucleus, green: CD3, red: CD4. ( I ) Percentage of CD4+CD8+T cells in brain tumors (* p < 0.05, ** p < 0.01). ( J , K ) Th17/Treg cells in brain tumor.

Article Snippet: The slices were treated in 3% hydrogen peroxide for 15 min at room temperature, washed with PBS for three times, incubated with normal goat serum at room temperature, and then incubated with rabbit anti-mouse CD4 antibody and rabbit anti-mouse CD8 primary (Bioss, Beijing, China; 1:100, 4 °C overnight) antibodies.

Techniques: Immunofluorescence, Immunohistochemical staining, Staining, Enzyme-linked Immunosorbent Assay

Journal: Pharmaceuticals

Article Title: AHR Agonist ITE Boosted PD1 Antibody’s Effects by Inhibiting Myeloid-Derived Cells Suppressive Cells in an Orthotopic Mouse Glioma Model

doi: 10.3390/ph18040471

Figure Lengend Snippet: Primary antibody list.

Article Snippet: The slices were treated in 3% hydrogen peroxide for 15 min at room temperature, washed with PBS for three times, incubated with normal goat serum at room temperature, and then incubated with rabbit anti-mouse CD4 antibody and rabbit anti-mouse CD8 primary (Bioss, Beijing, China; 1:100, 4 °C overnight) antibodies.

Techniques:

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Proportions of CD3+, CD8 +, and CD4 + T cells by flow cytometry in control and aCD40 treated Brpkp110 tumors on day 13 post treatment ( n = 10). B FoxP3 + CD4+ regulatory T cells on day 7 post-treatment ( n = 7). C Proportions of Granzyme B + T cells in subpopulations by flow cytometry in control and aCD40 treated Brpkp110 tumors on day 7 post implantation ( n = 10). D Images of immunofluorescent staining for CD8 (red) and nuclei (blue) and CD8 staining quantification in untreated (control) and aCD40 treated Brpkp110 tumors on day 7 post treatment ( n = 4–6). E Quantification of CD8 immunofluorescent staining in outer, middle, and inner thirds of control and aCD40 treated tumors on day 7 post treatment ( n = 5–6). F After 7 days of treatment with aCD40, Brpkp110 tumors were minced and cultured ex vivo. Supernatant was collected and pooled for each treatment group after 48 h and cytokines were measured ( n = 2, with 3 tumors pooled per group). G T cell activation and proliferation markers measured by flow cytometry in CD4+ and CD8 + T cell populations of control and aCD40 treated Brpkp110 TDLN on day 7 post-treatment ( n = 10). H Growth curves of control and aCD40 treated Brpkp110 tumors implanted into WT hosts with or without T cell depletions ( n = 17–20). I Tumor growth curve of aCD40 (treatment on Day 8) +/− FTY720 (treatment started on Day 7) treated Brpkp110 tumors ( n = 14–16). Data: ( A – C , D (right), E – G ) median, ( H , I ) mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Flow Cytometry, Control, Staining, Cell Culture, Ex Vivo, Activation Assay

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 26 post implantation (right) of Brpkp110 tumors. Indicated treatments initiated on day 7 post implantation ( n = 18–20). B Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 26 post implantation (right) of E0771 tumors. Indicated treatments initiated on day 8 post implantation ( n = 12–15). C Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 31 post implantation (right) of AT3 tumors. Indicated treatments initiated on day 9 post implantation ( n = 14). D Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 34 post implantation (right) of EpH4 1424 tumors. Indicated treatments initiated on day 5 post implantation ( n = 14–16). E Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 27 post implantation in control and aCD40+ICB treated hosts with and without CD8 + T cell depletions. Indicated treatments initiated on day 7 post implantation ( n = 12–15). F Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 25 post implantation in control and aCD40+ICB treated hosts with and without CD4 + T cell depletions. Indicated treatments initiated on day 8 post implantation ( n = 14–18). G Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 27 post implantation in control and aCD40+ICB treated hosts with and without CD4+ and CD8 + T cell depletions. Indicated treatments initiated on day 7 post implantation ( n = 12–18). H Tumor growth curves (left) and volume changes compared to pretreatment (right) on day 26 post Brpkp110 tumor implantation into WT and BATF3 KO hosts. Indicated treatments initiated on day 7 post implantation ( n = 14–18). Data: ( A – H left) mean ± SEM, ( A – H right) each column represents individual tumor. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Control, Tumor Implantation

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Proportions of circulating effector memory (CD44 + CD62L-), central memory (CD44 + CD62L+), and naïve (CD44-CD62L-) CD4+ (left) and CD8+ (right) in blood, 3 months post treatment induced tumor clearance ( n = 5–6, data representative of 2 experiments with similar results). B Secondary Brpkp110 tumor rechallenge of naïve and previously Brpkp110 tumor-bearing mice cured after aCD40 + ICB, at least 2 months post primary tumor clearance ( n = 12–14, data representative of 3 experiments with similar results). C Control and rechallenge tumor growth in T cell sufficient ( n = 6–12) and T cell depleted hosts ( n = 12–14, data representative of 2 experiments with similar results). D Brpkp110 tumor growth curves in intra-tumoral (IT) vehicle (control) and IT aCD40 treated hosts. aCD40 administered tumors denoted as aCD40 IT and contralateral untreated tumors denoted as CD40 IT Distant ( n = 9–12, data representative of 2 experiments with similar results). E Brpkp110 tumor growth curves in intra-tumoral (IT) vehicle (control) and IT or intraperitoneal (IP) aCD40 or ICB received hosts ( n = 4–9, data representative of 2 experiments with similar results). Data: ( A ) median, ( C – E ) mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Control

Journal: NPJ Vaccines

Article Title: Intradermal vaccination of live attenuated influenza vaccine protects mice against homologous and heterologous influenza challenges

doi: 10.1038/s41541-021-00359-8

Figure Lengend Snippet: Mice were challenged with 10x LD 50 of mouse-adapted A/Hong Kong/415742Md virus at 14 days after i.d. vaccination. Bronchoalveolar lavage fluid (BAL), lung, spleen, and serum samples were collected at 2 and 4 dpi. a T cell composition in mouse lung single-cell suspension by flow cytometry assays. Effector memory (EM) T cells were identified as CD3 + , CD4 + /CD8 + , CD44 + , and CD62L − cells, whereas central memory (CM) T cells were identified as CD3 + , CD4 + /CD8 + , CD44 + , and CD62L + cells. n = 3–5 mice. Error bars indicate standard error of mean. * p < 0.05; ** p < 0.01; *** p < 0.001 when compared with PBS group by two-way ANOVA. b CD4 + and CD8 + T cell distribution in lung sections at 2 dpi. Representative images of immunohistochemically stained NP, CD4, and CD8. In vaccinated mouse lung, some viral NP-expressing cells were observed in the bronchiolar epithelium (solid arrows), extensive CD4 + and CD8 + T cell infiltration around the same bronchiole (open arrows). In PBS control mouse lung, abundant viral antigen-expressing cells in the entire epithelium layer (solid arrows) while CD4 + and CD8 + T cell infiltration were much less frequent (open arrows). Scale bars = 100 µm. c Relative expression levels of IL-2, IL-4, and IFN-γ in lung at 2 dpi by real-time RT-PCR comparing to uninfected lung samples. Viral-specific interferon-γ producing CD4 + and CD8 + T cells in ( d ) BAL and spleen ( e ) taken at 4 dpi. In vitro T cell epitope stimulation with CD4 epitope and CD8 T epitope from influenza NP (CD4 T epitope peptide: RLIQNSITIERMVLS; CD8 epitope peptide: TYQRTRALV) were performed for 48 h. Right-hand side is the representative images from EISPOT assay. n = 3 per group. Error bars indicate standard error of mean. * p < 0.05; ** p < 0.01 when compared with PBS group by two-way ANOVA. f Antibody-mediated T cell depletion in vaccinated mice did not compromise vaccine-induced protection against A/Hong Kong/415742Md challenge, no weight loss (left) or lethality (right) were observed after virus challenge. g Representative images of H&E (left) and immunohistochemistry stained viral NP in the T cell-depleted mouse lung at 4 dpi. Scale bars = 100 µm. h Immunohistochemistry stained CD4, CD8 T cells, respectively, in the T cell-depleted or not depleted mouse lungs at 4 dpi. Scale bars = 100 µm. i Serum antibody titer, HI (left), MN (right) tested at 4 dpi in T cell-depleted or not depleted mice, and control mice. n = 3–5 per group.

Article Snippet: After blocking with 1% bovine serum albumin, the sections were incubated with mouse anti-influenza NP, rabbit anti-mouse CD4, or rabbit anti-mouse CD8 primary antibody (all from Abcam) at 4 °C overnight , followed by biotin-conjugated secondary antibody (Calbiochem, Darmstadt, Germany) for 30 min at room temperature.

Techniques: Virus, Suspension, Flow Cytometry, Staining, Expressing, Control, Quantitative RT-PCR, In Vitro, Immunohistochemistry