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MINUCELLS AND MINUTISSUE VERTRIEBS GMBH

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PAGE 16

Flow chart of cell detection on
non-transparent supports:


 

Detection of cells grown on non-transparent supports: Determination of cell growth characteristics on fully transparent supports can easily be obtained by conventional phase contrast microscopy. However, with semi- and non-transparent supports the conventional methods to determine the appropriate parameters fail. For that reason we suggest a quick and practical approach for detection of cells on non-transparent supports with an epifluorescence microscope. See references in proceedings.

 

Procedure takes 60 minutes or less:

1. For example, cells are cultured on the upper side of a support mounted on a tissue carrier so that the white tension ring can be seen.

2. Place the tissue carrier in a 24 well plate.

3. Fix the specimens in 70% ethanol for 10 minutes.

4. Stain for example in fluorescent DAPI or propidium iodide (4 mg/ml) in phosphate buffered saline without Ca2+ and Mg2+ for 5 respectively 30 minutes.

5. Wash 3 x 10 minutes in phosphate buffered saline.

6. Turn the tissue carrier upside down with fine forceps so that the cells are now on the lower side of the support within a 24 well culture plate. The white tension ring now faces the bottom of the plate.

7. Record the nuclear staining profile of the cells within phosphate buffered saline under the inverse epifluorescence microscope in a 24 well culture plate.

MDCK cells on optimal and non-optimal supports
MDCK cells on optimal and non-optimal supports

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