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Griffin:siRNA transfection: Difference between revisions
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→Suspension Cells
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===Suspension Cells=== | ===Suspension Cells=== | ||
Suspension cells require slow speed centrifugation (2500xg) to collect intact cell pellets from cuture flasks, then counting the cells with a hemocytometer and then seeding the wells accordingly by cell count into transfection medium. Suspension cells can proceed directly to the siRNA experiment rather than adherent cells, that require seeding the cells and waiting for adherence. | Suspension cells require slow speed centrifugation (2500xg) to collect intact cell pellets from cuture flasks, then counting the cells with a hemocytometer and then seeding the wells accordingly by cell count into transfection medium. Suspension cells can proceed directly to the siRNA experiment rather than adherent cells, that require seeding the cells and waiting for adherence. | ||
====Sorting Dead Suspension Cells===== | |||
Dead Cells are generally lighter than live cells. Resuspend the cells in 15 or 50 ml conical centrifuge tubes and allow the cells to settle 2-5 minutes. Most live cells will sediment to the bottom, leaving a mixture of Live/Dead cells floating in the supernatant. Aspirate supernatant and discard. This will leave the live cells ready for resuspension into the culture flask. Some live cells will be lost in this process. | |||
===Six well plates (2x10e5 cells per well)=== | ===Six well plates (2x10e5 cells per well)=== | ||
