DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
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DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay : results from the hCOMET ring trial. / Møller, Peter; Azqueta, Amaya; Rodriguez-Garraus, Adriana; Bakuradze, Tamara; Richling, Elke; Bankoglu, Ezgi Eyluel; Stopper, Helga; Bastos, Victoria Claudino; Langie, Sabine A S; Jensen, Annie; Ristori, Sara; Scavone, Francesca; Giovannelli, Lisa; Wojewódzka, Maria; Kruszewski, Marcin; Valdiglesias, Vanessa; Laffon, Blanca; Costa, Carla; Costa, Solange; Teixeira, João Paulo; Marino, Mirko; Del Bo', Cristian; Riso, Patrizia; Zhang, Congying; Shaposhnikov, Sergey; Collins, Andrew.
In: Mutagenesis, Vol. 38, No. 5, 2023, p. 273–282.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay
T2 - results from the hCOMET ring trial
AU - Møller, Peter
AU - Azqueta, Amaya
AU - Rodriguez-Garraus, Adriana
AU - Bakuradze, Tamara
AU - Richling, Elke
AU - Bankoglu, Ezgi Eyluel
AU - Stopper, Helga
AU - Bastos, Victoria Claudino
AU - Langie, Sabine A S
AU - Jensen, Annie
AU - Ristori, Sara
AU - Scavone, Francesca
AU - Giovannelli, Lisa
AU - Wojewódzka, Maria
AU - Kruszewski, Marcin
AU - Valdiglesias, Vanessa
AU - Laffon, Blanca
AU - Costa, Carla
AU - Costa, Solange
AU - Teixeira, João Paulo
AU - Marino, Mirko
AU - Del Bo', Cristian
AU - Riso, Patrizia
AU - Zhang, Congying
AU - Shaposhnikov, Sergey
AU - Collins, Andrew
N1 - © The Author(s) 2023. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
PY - 2023
Y1 - 2023
N2 - The comet assay is widely used in biomonitoring studies for the analysis of DNA damage in leukocytes and peripheral blood mononuclear cells. Rather than processing blood samples directly, it can be desirable to cryopreserve whole blood or isolated cells for later analysis by the comet assay. However, this creates concern about artificial accumulation of DNA damage during cryopreservation. In this study, ten laboratories used standardized cryopreservation and thawing procedures of monocytic (THP-1) or lymphocytic (TK6) cells. Samples were cryopreserved in small aliquots in 50% foetal bovine serum, 40% cell culture medium and 10% dimethyl sulphoxide. Subsequently, cryopreserved samples were analysed by the standard comet assay on three occasions over a three-year period. Levels of DNA strand breaks in THP-1 cells were increased (4 laboratories), unaltered (4 laboratories) or decreased (2 laboratories) by long-term storage. Pooled analysis indicates only a modest positive association between storage time and levels of DNA strand breaks in THP-1 cells (0.37% Tail DNA per year, 95% confidence interval: -0.05, 0.78). In contrast, DNA strand break levels were not increased by cryopreservation in TK6 cells. There was inter-laboratory variation in levels of DNA strand breaks in THP-1 cells (SD = 3.7% Tail DNA) and TK6 reference sample cells (SD = 9.4% Tail DNA), whereas the intra-laboratory residual variation was substantially smaller (i.e. SD = 0.4% to 2.2% Tail DNA in laboratories with the smallest and largest variation). In conclusion, the study shows that accumulation of DNA strand breaks in cryopreserved mononuclear blood cell lines is not a matter of concern.
AB - The comet assay is widely used in biomonitoring studies for the analysis of DNA damage in leukocytes and peripheral blood mononuclear cells. Rather than processing blood samples directly, it can be desirable to cryopreserve whole blood or isolated cells for later analysis by the comet assay. However, this creates concern about artificial accumulation of DNA damage during cryopreservation. In this study, ten laboratories used standardized cryopreservation and thawing procedures of monocytic (THP-1) or lymphocytic (TK6) cells. Samples were cryopreserved in small aliquots in 50% foetal bovine serum, 40% cell culture medium and 10% dimethyl sulphoxide. Subsequently, cryopreserved samples were analysed by the standard comet assay on three occasions over a three-year period. Levels of DNA strand breaks in THP-1 cells were increased (4 laboratories), unaltered (4 laboratories) or decreased (2 laboratories) by long-term storage. Pooled analysis indicates only a modest positive association between storage time and levels of DNA strand breaks in THP-1 cells (0.37% Tail DNA per year, 95% confidence interval: -0.05, 0.78). In contrast, DNA strand break levels were not increased by cryopreservation in TK6 cells. There was inter-laboratory variation in levels of DNA strand breaks in THP-1 cells (SD = 3.7% Tail DNA) and TK6 reference sample cells (SD = 9.4% Tail DNA), whereas the intra-laboratory residual variation was substantially smaller (i.e. SD = 0.4% to 2.2% Tail DNA in laboratories with the smallest and largest variation). In conclusion, the study shows that accumulation of DNA strand breaks in cryopreserved mononuclear blood cell lines is not a matter of concern.
U2 - 10.1093/mutage/gead019
DO - 10.1093/mutage/gead019
M3 - Journal article
C2 - 37357800
VL - 38
SP - 273
EP - 282
JO - Mutagenesis
JF - Mutagenesis
SN - 0267-8357
IS - 5
ER -
ID: 360133055