Programm                 "Degeneration und Regeneration– Grundlagen, Diagnostik und Therapie"

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Impression Cytology and Analysis by Flow Cytometry: Influence of Culture Medium, Storage Temperature and Period on Cellular Output

Repp A., Dick H. B., Pfeiffer N., Grus F. H.
Dept. of Ophthalmology, Mainz University

Purpose: Analysis of impression cytology specimens allows examination of surface and intracellular antigens. The purpose of this study was to assess optimal experimental conditions for acquisition and analysis to separate a homogeneous population of cells.
Method: Specimens were collected from nonexposed conjunctiva of healthy patients. Cellulose acetate filters were applied on the superior and superotemporal bulbar conjunctiva. The membranes were immediately dipped into tubes containing ethanol 70%, paraformaldehyde 0,05%, and bovines serum albumin 1%. Tubes were kept at +4°C, -20°C and -80°C. The conjunctival samples were processed up to 8 hours, 3,7, and 14 days after collection.
Results: The linear plot giving granulometry versus cell size revealed a single cell population. Exclusion of cellular debris, aggregates, and background fluorescence was most successful in tubes kept in bovine serum albumin at +4°C for 8 hours. Long-term storage and lower culture temperature led to sample degradation and loss of cellular output.
Conclusions: Flow cytometry allows identification of single cell populations after impression cytology. Culture of samples in bovine serum albumin 1% at 4° showed a homogenous single cell population. Storage and processing times have to be kept as short as possible.

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