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Found 3 matching solutions for this experiment
| Upstream tips |
Protocol tips |
Downstream tips |
If you are unfamiliar with cell type being used, culture an extra dish of cells for determining cell number.
-Before crosslinking, trypsinize and determine the cell number from the extra dish of cells. |
-After incubation, it is essential to open the column cap before plug removal to equalize the pressure within the column. Failure to open the cap before plug removal will result in sample loss. |
-If you are preparing chromatin in bulk, unused supernatant may be stored at -80°C for later use. |
| Upstream tips |
If you are unfamiliar with cell type being used, culture an extra dish of cells for determining cell number.
-Before crosslinking, trypsinize and determine the cell number from the extra dish of cells. |
| Protocol tips |
| -After incubation, it is essential to open the column cap before plug removal to equalize the pressure within the column. Failure to open the cap before plug removal will result in sample loss. |
| Downstream tips |
| -If you are preparing chromatin in bulk, unused supernatant may be stored at -80°C for later use. |
| Upstream tips |
Protocol tips |
Downstream tips |
-The 1.25 M glycine must be at room temperature before use.
-The Lysis Buffer must be at room temperature and fully resuspended before use. Vortex briefly to resuspend. |
-For each ChIP reaction, use 10,000–300,000 cells or 0.167–5 mg of tissue.
-1–10 μg of antibody is a typical starting range.
- 10-minute crosslinking step using formaldehyde at a 1% final concentration.
-Keep the cell lysate cooled on ice during sonication. |
|
| Upstream tips |
-The 1.25 M glycine must be at room temperature before use.
-The Lysis Buffer must be at room temperature and fully resuspended before use. Vortex briefly to resuspend. |
| Protocol tips |
-For each ChIP reaction, use 10,000–300,000 cells or 0.167–5 mg of tissue.
-1–10 μg of antibody is a typical starting range.
- 10-minute crosslinking step using formaldehyde at a 1% final concentration.
-Keep the cell lysate cooled on ice during sonication. |
| Upstream tips |
Protocol tips |
Downstream tips |
|
-Just because an antibody works well in a Western blot does not always indicate it will perform well in chromatin immunoprecipitation.
Unlike a Western blot that detects proteins that have been denatured, a ChIP antibody must recognize the target protein in its native state. Use ChIP-validated antibody.
-Use 2-10 μg of your ChIP antibody depending on the abundance of your protein target. |
|
| Protocol tips |
-Just because an antibody works well in a Western blot does not always indicate it will perform well in chromatin immunoprecipitation.
Unlike a Western blot that detects proteins that have been denatured, a ChIP antibody must recognize the target protein in its native state. Use ChIP-validated antibody.
-Use 2-10 μg of your ChIP antibody depending on the abundance of your protein target. |
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