FACS analysis of CD4+ and CD8+ T cells infiltrating tumor tissues was performed using mouse anti-CD4+ FITC (clone GK1.5) and mouse anti-CD8+ FITC (clone 53–6.7), (BD Pharmingen, San Diego, CA) . Tumor cells were suspended in PBS at a concentration of 1 × 106 cell/ml. Cells were prepared as described above before being centrifuged. The supernatant was discarded. Cell pellets were then re-suspended in 500 μl PBS before 1 ml of suspension was dispensed in flow cytometric tube. Cells were incubated with 25 μl of anti-CD4+ or anti-CD8 in dark for 30 min at 4 °C. After the supernatant was discarded, the cells were washed twice by PBS, pH 7.2. Two hundred microliter paraformaldehyde solution was then added to each tube, mixed well, and kept in dark at 4 °C till FACS analysis was conducted according to the manufacturer′s instructions.
Mouse anti cd4 fitc clone gk1.5
Manufactured by BD
Mouse anti-CD4+ FITC (clone GK1.5) is a fluorescently-labeled monoclonal antibody that binds to the CD4 cell surface marker. CD4 is a glycoprotein expressed on the surface of T helper cells, regulatory T cells, and monocytes. This product can be used for the identification and enumeration of CD4+ cells in flow cytometry applications.
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2 protocols using mouse anti cd4 fitc clone gk1.5
1
Flow Cytometric Analysis of T-Cell Infiltration
2
Flow Cytometric Analysis of Tumor-Infiltrating T Cells
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Flow cytometric analysis (FACS) of CD4+ and CD8+ T cells that infiltrated EAC tumors was performed using mouse anti-CD4+ FITC (clone GK1.5) and mouse anti-CD8+ FITC (clone 53–6.7) (BD Biosciences). Tumor cell suspensions in PBS at a concentration of 1×106 cells/ml were prepared, centrifuged for 5 min at 200–300 × g, and the supernatant was discarded. Afterwards the cell pellets were re-suspended in 500 µl PBS. Subsequently, 1 ml of suspension was dispensed in a flow cytometric tube. Cells were incubated with 25 µl of anti-CD4+ or anti-CD8+ in the dark for 30 min at 4°C. The supernatant was discarded and the cells were washed twice by PBS, pH (7.2). Then, 200 µl paraformaldehyde solution was added to each tube, mixed well and maintained in the dark at 4°C until FACS analysis following the manufacturers instructions.
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