CY5-anti-CD3, FITC-anti-CD8, CY5-anti-CD8, CY5-anti-CD4, PE-anti-CD158b (KIR2DS2), PE-anti-NKB1 (KIR3DL1) and FITC-anti-CD28 were obtained from BD PharMingen, San Diego, CA. PE-anti-CD158d (KIR2DL4) was obtained from R&D Systems (Minneapolis, MN) and PE-anti-CD158a/h (KIR2DL1/2DS2) and PE-anti-CD158b1/b2 (KIR 2DL2/2DL3/2DS2) were obtained from Beckman Coulter Immunotech (Fullerton, CA). Five μl of each PE-conjugated anti-KIR Ab were mixed to form a “cocktail” and used to stain the cells (19 (link)). All labeling procedures were performed on ice in PBS containing 10% horse serum and normal human AB serum (Invitrogen) and sodium azide. Cell staining was performed as previously described (20 (link)), and the stained cells were analyzed using a FACSCalibur flow cytometer (BD Biosciences, Franklin Lakes, NJ).
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Multicolor Flow Cytometry Analysis
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Other organizations : University of Michigan–Ann Arbor
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Variable analysis
independent variables
- CY5-anti-CD3
- FITC-anti-CD8
- CY5-anti-CD8
- CY5-anti-CD4
- PE-anti-CD158b (KIR2DS2)
- PE-anti-NKB1 (KIR3DL1)
- FITC-anti-CD28
- PE-anti-CD158d (KIR2DL4)
- PE-anti-CD158a/h (KIR2DL1/2DS2)
- PE-anti-CD158b1/b2 (KIR 2DL2/2DL3/2DS2)
- Staining of cells
- PBS containing 10% horse serum and normal human AB serum (Invitrogen) and sodium azide
- Cell staining procedure as previously described
- Not specified
- Not specified
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