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Found 3 matching solutions for this experiment
| Upstream tips |
Protocol tips |
Downstream tips |
-Warm IP Lysis Buffer to room temperature to prevent precipitation.
-Prepare one extra plate of cells to estimate cell number.
-Thaw Protease Inhibitor Cocktail (PIC) at room temperature.
-It is recommended to take some trial to optimize sonication condition before beginning with sonication. |
-In general it is recommended that one million mammalian cells are required for each IP
fraction.
-Centrifuge the sample at 6000 × g for 5 min at 4 °C to eliminate bubbles.
-To reduce background, remove as much supernatant as possible in the last wash. Continue with next step OR store at 4°C overnight. |
|
| Upstream tips |
-Warm IP Lysis Buffer to room temperature to prevent precipitation.
-Prepare one extra plate of cells to estimate cell number.
-Thaw Protease Inhibitor Cocktail (PIC) at room temperature.
-It is recommended to take some trial to optimize sonication condition before beginning with sonication. |
| Protocol tips |
-In general it is recommended that one million mammalian cells are required for each IP
fraction.
-Centrifuge the sample at 6000 × g for 5 min at 4 °C to eliminate bubbles.
-To reduce background, remove as much supernatant as possible in the last wash. Continue with next step OR store at 4°C overnight. |
| 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 |
| -Do not re-freeze the Protein G magnetic beads, the beads should be stored at 4C. |
-Always use ChIP validated antibody.
-Optimal use of 1-3 ug of antibody. |
-The diluted Proteinase K stop solution can not be stored. |
| Upstream tips |
| -Do not re-freeze the Protein G magnetic beads, the beads should be stored at 4C. |
| Protocol tips |
-Always use ChIP validated antibody.
-Optimal use of 1-3 ug of antibody. |
| Downstream tips |
| -The diluted Proteinase K stop solution can not be stored. |
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