Short/Long-Term Cryopreservation Prior To Comet Assay Of Whole-Blood Leukocytes And In Vitro-Cultured Lung Fibroblasts

Single-cell gel electrophoresis (comet assay) is a valuable test that can be used in ecotoxicological, epidemiological, and biomonitoring contexts. We assessed the effects of short- (without cryopreservation) and long-term (with cryopreservation) storage of DMEM-cultivated human peripheral blood leukocytes (HPBLs) and a human lung fibroblast cell line (FLECH-104) on comet assay results. Samples were stored for 6 or 24 h at room temperature (23 degrees C) or 4 degrees C and frozen at -80 degrees C or -196 degrees C for 1, 2, or 4 weeks. Short-term storage led to significant increases in the comet tail intensity (TI) and Olive tail moment (OTM) in HPBL and FLECH-104 samples. Freezing FLECH-104 samples at -80 degrees C and -196 degrees C resulted in TI mean increases, with no differences in OTM. All frozen HPBL samples did not exhibit significant increases in TI or OTM, and instead exhibited a slight decrease in TI versus the control at both -80 degrees C and -196 degrees C. Increased frequency of highly damaged cells was observed in FLECH-104 and HPBL cultures during both short-term storage and after freezing, which may indicate a significant destructive effect. Therefore, freezing of cell cultures and whole blood according to our protocol is not recommended.