[Preservation of mucosal specimens before processing in the alkaline single cell microgel electrophoresis assay].
BACKGROUND
Human mucosal biopsies are established in ecogenotoxicological studies, but up until now they have demanded immediate processing after harvesting. We report our experience with the preservation of specimens either for 24 h at 4 degrees C or for longer periods at -80 degrees C and compare the results with fresh specimens using the alkaline single cell microgel electrophoresis assay.


METHODS
Nasal mucosa was harvested from ten patients, transferred to the laboratory and divided into groups for immediate processing,24 h preservation at 4 degrees C and cryopreservation at -80 degrees C. Alkaline single cell microgel electrophoresis assays were performed after separating the specimens into single cells and after exposure to benzo[a]pyrene,benzo[a]pyrene-diolepoxide, N-nitrosodiethylamine, or sodium dichromate. The trypan blue exclusion test was used to assess cytotoxicity.


RESULTS
Despite of the fact that cell viability remained stable, after cryopreservation DNA-migration increased significantly for the negative control and benzo[a]pyrene. Although an increase was also seen for sodium dichromate, this was not significant. For benzo[a]pyrene-diolepoxide, N-nitrosodiethylamine and N-methyl-N'-nitro-N-nitrosoguanidine there were no significant changes in DNA-migration. After 24 h in cell medium at 4 degrees C,DNA-migration did not rise compared to the samples which were immediately processed.


CONCLUSIONS
Preservation of mucosal specimens at 4 degrees C for 24 h may be legitimate in order to facilitate laboratory practice. However, cryopreservation should not be applied because it leads to higher rates of DNA migration in some tested substances in the alkaline single cell microgel electrophoresis assay.
Kleinsasser
N H
NH
Klinisch experimentelle Onkologie der Klinik und Poliklinik für Hals-, Nasen- und Ohrenkranke der Ludwig-Maximilians-Universität München. norbert.kleinsasser@klinik.uni-regensburg.de
Kastenbauer
E R
ER
Zieger
S
S
Baluschko
T
T
Wallner
B C
BC
Harréus
U A
UA
D000293:Adolescent; D000328:Adult; D001706:Biopsy; D015197:Carcinogenicity Tests; D002274:Carcinogens, Environmental / Q000633:toxicity; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D020552:Comet Assay; D015925:Cryopreservation; D004249:DNA Damage; D004784:Environmental Monitoring; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009297:Nasal Mucosa / Q000187:drug effects / Q000473:pathology*; D013048:Specimen Handling; D020858:Tissue and Organ Harvesting
D002274:Carcinogens, Environmental