Counting cells with a hemocytometer.
- Tube of cell suspension
- Trypan Blue 0.4%
- Combine 20μl cell suspension with 20μl of Trypan Blue. Mix.
- Load 10μl into each of the 2 hemocytometer "sides" using the capillary action created by the coverslip and the hemocytometer. Avoid bubbles.
- Count the number of cells in 4 corners of the gridded area of each side. (See below) White cells are live and blue cells are dead/dying.
- Record cell count and repeat for the other side of the hemocytometer.
- If the two cell counts are roughly equivalent, average the cell counts. Otherwise, recount.
- Mean cell count/4 * Dilution = Cells x 104/ml.
- Above example, Mean cell count/4 * 2 = Cells x 104/ml.
- Average number of 3T12 cells on a confluent:
- T175: 35-50M (LSJ, N=2)
- 10cm dish: 3M (VT)
- Cell doubling time:
- 3T12: ~24hrs.