The most common harvest method is by trypsinization. Use what ever method that you feel yields the best results. Once a good quality cell suspension has been established then stain the cells using the method outlined above. Adherent cell lines often do not lyse as completely as non adherent cell lines, leaving part of the RNA rich cytoplasm attached to the nucleus. It is very important that a high quality suspension of stained, single nuclei be established before attempting analysis on the flow cytometer.
Some cell lines can be very difficult to prepare successfully.
Therefore, it is highly recommended that nuclei suspensions be examined under a fluorescent microscope to determine their quality before running them on the flow cytometer. The Flow Cytometry Facility has a fluorescent microscope available for this purpose. An alternative harvest method to trypsinization is to add hypotonic PI solution directly to the cell flask. A reasonable estimation of the cell numbers in the flask should be determined first so that the appropriate volume of hypotonic PI solution can be added.