DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial

Detalhes bibliográficos
Autor(a) principal: Møller, Peter
Data de Publicação: 2023
Outros Autores: Azqueta, Amaya, Rodriguez-Garraus, Adriana, Bakuradze, Tamara, Richling, Elke, Bankoglu, Ezgi Eyluel, Stopper, Helga, Claudino Bastos, Victoria, Langie, Sabine A.S., Jensen, Annie, Ristori, Sara, Scavone, Francesca, Giovannelli, Lisa, Wojewódzka, Maria, Kruszewski, Marcin, Valdiglesias, Vanessa, Laffon, Blanca, Costa, Carla, Costa, Solange, Paulo Teixeira, João, Marino, Mirko, Del Bo, Cristian, Riso, Patrizia, Zheng, Congying, Shaposhnikov, Sergey, Collins, Andrew
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10400.18/9050
Resumo: 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, 10 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 3-year period. Levels of DNA strand breaks in THP-1 cells were increased (four laboratories), unaltered (four laboratories), or decreased (two 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%-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.
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spelling DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trialComet AssayDNA Strand BreaksCryopreservationGenotoxicityValidationGenotoxicidade AmbientalThe 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, 10 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 3-year period. Levels of DNA strand breaks in THP-1 cells were increased (four laboratories), unaltered (four laboratories), or decreased (two 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%-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.Certain authors have acknowledged funding as follows: statutory funding for INTC and IRH (M.W. and M.K.); Spanish Ministry of Science and Innovation MCIN/ AEI/10.13039/501100011033 (Grants PID2020- 113788RB-I00 and PID2020-114908 GA-I00), and Xunta de Galicia (Grant ED431B 2022/16) (V.V. and B.L.); Università degli Studi di Firenze fund RICATEN21 and RICATEN22 (L.G.).Oxford University Press/ UK Environmental Mutagen SocietyRepositório Científico do Instituto Nacional de SaúdeMøller, PeterAzqueta, AmayaRodriguez-Garraus, AdrianaBakuradze, TamaraRichling, ElkeBankoglu, Ezgi EyluelStopper, HelgaClaudino Bastos, VictoriaLangie, Sabine A.S.Jensen, AnnieRistori, SaraScavone, FrancescaGiovannelli, LisaWojewódzka, MariaKruszewski, MarcinValdiglesias, VanessaLaffon, BlancaCosta, CarlaCosta, SolangePaulo Teixeira, JoãoMarino, MirkoDel Bo, CristianRiso, PatriziaZheng, CongyingShaposhnikov, SergeyCollins, Andrew2024-02-08T10:39:53Z2023-10-142023-10-14T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10400.18/9050engMutagenesis. 2023 Oct 14;38(5):273-282. doi: 10.1093/mutage/gead0190267-835710.1093/mutage/gead019metadata only accessinfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-02-10T01:30:46Zoai:repositorio.insa.pt:10400.18/9050Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T02:37:21.661377Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse
dc.title.none.fl_str_mv DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
title DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
spellingShingle DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
Møller, Peter
Comet Assay
DNA Strand Breaks
Cryopreservation
Genotoxicity
Validation
Genotoxicidade Ambiental
title_short DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
title_full DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
title_fullStr DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
title_full_unstemmed DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
title_sort DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial
author Møller, Peter
author_facet Møller, Peter
Azqueta, Amaya
Rodriguez-Garraus, Adriana
Bakuradze, Tamara
Richling, Elke
Bankoglu, Ezgi Eyluel
Stopper, Helga
Claudino Bastos, Victoria
Langie, Sabine A.S.
Jensen, Annie
Ristori, Sara
Scavone, Francesca
Giovannelli, Lisa
Wojewódzka, Maria
Kruszewski, Marcin
Valdiglesias, Vanessa
Laffon, Blanca
Costa, Carla
Costa, Solange
Paulo Teixeira, João
Marino, Mirko
Del Bo, Cristian
Riso, Patrizia
Zheng, Congying
Shaposhnikov, Sergey
Collins, Andrew
author_role author
author2 Azqueta, Amaya
Rodriguez-Garraus, Adriana
Bakuradze, Tamara
Richling, Elke
Bankoglu, Ezgi Eyluel
Stopper, Helga
Claudino Bastos, Victoria
Langie, Sabine A.S.
Jensen, Annie
Ristori, Sara
Scavone, Francesca
Giovannelli, Lisa
Wojewódzka, Maria
Kruszewski, Marcin
Valdiglesias, Vanessa
Laffon, Blanca
Costa, Carla
Costa, Solange
Paulo Teixeira, João
Marino, Mirko
Del Bo, Cristian
Riso, Patrizia
Zheng, Congying
Shaposhnikov, Sergey
Collins, Andrew
author2_role author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Repositório Científico do Instituto Nacional de Saúde
dc.contributor.author.fl_str_mv Møller, Peter
Azqueta, Amaya
Rodriguez-Garraus, Adriana
Bakuradze, Tamara
Richling, Elke
Bankoglu, Ezgi Eyluel
Stopper, Helga
Claudino Bastos, Victoria
Langie, Sabine A.S.
Jensen, Annie
Ristori, Sara
Scavone, Francesca
Giovannelli, Lisa
Wojewódzka, Maria
Kruszewski, Marcin
Valdiglesias, Vanessa
Laffon, Blanca
Costa, Carla
Costa, Solange
Paulo Teixeira, João
Marino, Mirko
Del Bo, Cristian
Riso, Patrizia
Zheng, Congying
Shaposhnikov, Sergey
Collins, Andrew
dc.subject.por.fl_str_mv Comet Assay
DNA Strand Breaks
Cryopreservation
Genotoxicity
Validation
Genotoxicidade Ambiental
topic Comet Assay
DNA Strand Breaks
Cryopreservation
Genotoxicity
Validation
Genotoxicidade Ambiental
description 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, 10 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 3-year period. Levels of DNA strand breaks in THP-1 cells were increased (four laboratories), unaltered (four laboratories), or decreased (two 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%-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.
publishDate 2023
dc.date.none.fl_str_mv 2023-10-14
2023-10-14T00:00:00Z
2024-02-08T10:39:53Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
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format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10400.18/9050
url http://hdl.handle.net/10400.18/9050
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv Mutagenesis. 2023 Oct 14;38(5):273-282. doi: 10.1093/mutage/gead019
0267-8357
10.1093/mutage/gead019
dc.rights.driver.fl_str_mv metadata only access
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dc.publisher.none.fl_str_mv Oxford University Press/ UK Environmental Mutagen Society
publisher.none.fl_str_mv Oxford University Press/ UK Environmental Mutagen Society
dc.source.none.fl_str_mv reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
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