Cell Cultures - Eye Examination
The general health of any culture needs to be examined before an eye examination in cell cultures. This can be achieved quickly and quantitatively by the following observations:
- Examine the pH of the cell culture using the colour as an indicator. If the cell culture has become more acidic, the phenol red will shift from red to purple. Typically, cells can tolerate slight acidity better than basicity. A shift above pH 7.6 can be detrimental – colour standards can be used for comparison if unfamiliar with the various pH levels.
- The cell attachment of the monolayer cultures should be spread out evenly and to the well. Cells may be in a state of division or dying if they are floating in the culture. A dying cell will have irregular morphology.
- The growth rate of a culture is estimated by following it toward the development of a full cell sheet. When the amount of space covered by the cells is compared to the unoccupied spaces, the percent confluency can be estimated.
- An important guide is the shape of the cell as round cells in an uncrowded culture are not a good indicator unless they are dividing cells. Look for the doublets or dividing calls and know the effect of crowding on the cell shape.
- The number of ‘giant cells’ increases as the culture ages or declines in ‘well-being’. The frequency of giant cells should be low and constant under uniform culture conditions.
- A valuable early indicator is the success of a ‘culture split’ which is the rate at which the cells in the newly established cultures attach and spread out. An attachment within the first couple of hours suggests that the cells aren’t traumatised, and the in vitro environment is not abnormal. Good cell cultures can still occur even if an attachment doesn’t form in four hours.
- It is important to recognise the range of cell shapes and growth patterns shown by each cell line. Many transformed cells can ‘pile up’ due to a lack of contact inhibition and the effects of this are noticeable as the cells culture becomes overcrowded.