B16-F10 tumors were harvested on day 16–18 post tumor inoculation for flow cytometric analysis of in vivo experiments. Single cell suspensions were prepared using collagenase/hyaluronidase and DNase and red blood cells were lysed using ACK Lysis Buffer (Life Technologies). Live/Dead fixable yellow dead cell staining kit (Life Technologies) was applied for live/dead cell discrimination. Before surface staining, samples were incubated with Fc Block for 5 min on ice, followed by surface staining with anti-mouse CD45, CD3, CD8, CD4 (Supplementary Table 1 ). Cells were then fixed, permeabilized, and stained for intracellular FOXP3 (eBioscience). T effector cells were phenotyped as CD8+ and regulatory T cells (Treg cells) as CD4+FOXP3+. All flow cytometric analysis was done using a Beckman Coulter CyAn ADP and analyzed using software Summit 5.2. Flow cytometric data analysis was performed in a blinded fashion. Two independent researchers performed collection and analysis of flow cytometric data. Differences were compared and the overall P value was calculated by Mann Whitney test using the GraphPad Prism 5.0. (P value: *, P<0.05; **, P<0.01; ***, P<0.005.) The representative plots of relative abundance of tumor infiltrating T cells were showed in Supplementary Fig. 9 .
Live dead fixable yellow dead cell staining kit
Manufactured by Thermo Fisher Scientific
The Live/Dead Fixable Yellow Dead Cell Staining Kit is a fluorescent dye-based labeling solution used to detect and identify dead cells in a sample. The dye reacts with the amines in dead cells, providing a distinct signal that can be analyzed using flow cytometry or microscopy. This kit allows for the accurate detection and quantification of cell viability within a population.
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Lab products found in correlation
3 protocols using live dead fixable yellow dead cell staining kit
1
Quantifying Tumor-Infiltrating T Cells
2
Quantifying Tumor-Infiltrating T Cells
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B16-F10 tumors were harvested on day 16–18 post tumor inoculation for flow cytometric analysis of in vivo experiments. Single cell suspensions were prepared using collagenase/hyaluronidase and DNase and red blood cells were lysed using ACK Lysis Buffer (Life Technologies). Live/Dead fixable yellow dead cell staining kit (Life Technologies) was applied for live/dead cell discrimination. Before surface staining, samples were incubated with Fc Block for 5 min on ice, followed by surface staining with anti-mouse CD45, CD3, CD8, CD4 (Supplementary Table 1 ). Cells were then fixed, permeabilized, and stained for intracellular FOXP3 (eBioscience). T effector cells were phenotyped as CD8+ and regulatory T cells (Treg cells) as CD4+FOXP3+. All flow cytometric analysis was done using a Beckman Coulter CyAn ADP and analyzed using software Summit 5.2. Flow cytometric data analysis was performed in a blinded fashion. Two independent researchers performed collection and analysis of flow cytometric data. Differences were compared and the overall P value was calculated by Mann Whitney test using the GraphPad Prism 5.0. (P value: *, P<0.05; **, P<0.01; ***, P<0.005.) The representative plots of relative abundance of tumor infiltrating T cells were showed in Supplementary Fig. 9 .
3
Quantification of Tumor-Infiltrating T Cells
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Example 13
Tumors were harvested on day 16 post tumor inoculation for flow cytometric analysis of in vivo experiments. Single cell suspensions were prepared using collagenase/hyaluronidase and DNase and red blood cells were lysed using ACK Lysis Buffer (Life Technologies). Live/dead fixable yellow dead cell staining kit (Life Technologies) was applied for live/dead cell discrimination. Before surface staining, samples were incubated with Fc Block for 5 min on ice, followed by surface staining with anti-mouse CD45, CD3, CD8, CD4 (Table 1). Cells were then fixed, permeabilized, and stained for intracellular FOXP3 (eBioscience). T effector cells were phenotyped as CD8+ and regulatory T cells (Treg cells) as CD4+FOXP3+. All flow cytometric analysis was done using a Beckman Coulter CyAn ADP and analyzed using software Summit 5.2. Flow cytometric data analysis was performed in a blinded fashion. Two independent researchers performed collection and analysis of flow cytometric data. Differences were compared and the overall P value was calculated by analysis of unpaired t test using the GraphPad Prism 5.0. (P value: *, P<0.05; **, P<0.01; ***, P<0.005.) The representative plots of relative abundance of tumor infiltrating T cells were showed in
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