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Found 3 matching solutions for this experiment
| Upstream tips |
Protocol tips |
Downstream tips |
| Immortal macrophages, primary bone marrow derived macrophages and DCs or splenic DCs (0.5×106) of the indicated genotypes were treated with E.coli LPS and/or oxPAPC for the indicated time points at 37°C. |
Cells were then washed with 1 mL cold PBS and stained for appropriate antibodies on ice for 20 to 30 min. 2% mouse serum or rat serum were used as the blocking reagent to reduce non-specific binding of the antibodies. The stained cells were then washed with 1ml cold PBS and resuspended in 200 μL PBS. |
Staining of the surface receptors was analyzed with BD FACSCanto II. |
| Upstream tips |
| Immortal macrophages, primary bone marrow derived macrophages and DCs or splenic DCs (0.5×106) of the indicated genotypes were treated with E.coli LPS and/or oxPAPC for the indicated time points at 37°C. |
| Protocol tips |
| Cells were then washed with 1 mL cold PBS and stained for appropriate antibodies on ice for 20 to 30 min. 2% mouse serum or rat serum were used as the blocking reagent to reduce non-specific binding of the antibodies. The stained cells were then washed with 1ml cold PBS and resuspended in 200 μL PBS. |
| Downstream tips |
| Staining of the surface receptors was analyzed with BD FACSCanto II. |
| Upstream tips |
Protocol tips |
Downstream tips |
| Immortal macrophages, primary bone marrow derived macrophages and DCs or splenic DCs (0.5×106) of the indicated genotypes were treated with E.coli LPS and/or oxPAPC for the indicated time points at 37°C. |
Cells were then washed with 1 mL cold PBS and stained for appropriate antibodies on ice for 20 to 30 min. 2% mouse serum or rat serum were used as the blocking reagent to reduce non-specific binding of the antibodies. The stained cells were then washed with 1ml cold PBS and resuspended in 200 μL PBS. |
Staining of the surface receptors was analyzed with BD FACSCanto II. |
| Upstream tips |
| Immortal macrophages, primary bone marrow derived macrophages and DCs or splenic DCs (0.5×106) of the indicated genotypes were treated with E.coli LPS and/or oxPAPC for the indicated time points at 37°C. |
| Protocol tips |
| Cells were then washed with 1 mL cold PBS and stained for appropriate antibodies on ice for 20 to 30 min. 2% mouse serum or rat serum were used as the blocking reagent to reduce non-specific binding of the antibodies. The stained cells were then washed with 1ml cold PBS and resuspended in 200 μL PBS. |
| Downstream tips |
| Staining of the surface receptors was analyzed with BD FACSCanto II. |
| Upstream tips |
Protocol tips |
Downstream tips |
| Single-cell suspensions were incubated with a Fc receptor block (1 μg/1 × 106 cells; BD Bioscience, San Diego, CA) to reduce nonspecific anti-body binding. |
Dead cells were excluded using a Live/Dead Fixable Blue Dead Cell Stain Kit (Invitrogen) or 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma-Aldrich, St. Louis, MO). |
Flow cytometry was performed using BD LSR II and BD FACS Aria III flow cytometers (BD Bioscience), and data were analyzed with FlowJo software (TreeStar, Ashland, OR). |
| Upstream tips |
| Single-cell suspensions were incubated with a Fc receptor block (1 μg/1 × 106 cells; BD Bioscience, San Diego, CA) to reduce nonspecific anti-body binding. |
| Protocol tips |
| Dead cells were excluded using a Live/Dead Fixable Blue Dead Cell Stain Kit (Invitrogen) or 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma-Aldrich, St. Louis, MO). |
| Downstream tips |
| Flow cytometry was performed using BD LSR II and BD FACS Aria III flow cytometers (BD Bioscience), and data were analyzed with FlowJo software (TreeStar, Ashland, OR). |
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