Document Detail


Cryopreservation of human cumulus cells for co-cultures and assessment of DNA damage after thawing using the comet assay.
MedLine Citation:
PMID:  11699124     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
PURPOSE: Cumulus cells have been shown to be beneficial for blastocysts formation in co-cultures but information on cumulus cryopreservation is lacking. The objective was to use the fixed cell comet assay to analyze for DNA damage in cumulus cells after cryopreservation.
METHODS: Discarded cumulus cells from follicular aspirates obtained during assisted reproduction procedures (N = 4 cases) were pooled and cryopreserved in either 40% ethylene glycol and 0.5 M sucrose, 12:20% glycerol-egg yolk medium, 28% glycerol hypoosmolar medium or control medium. The cells were processed and stored in liquid nitrogen for 48 h. The thawed cells were smeared on glass slides, fixed, stained with acridine orange, embedded in a mini-agarose layer, and electrophoresis carried out. Fluorescent images were analyzed.
RESULTS: The cumulus tail moment, a calculated index of DNA damage, was significantly lower for each of the three cryoprotectant when compared with the control. The two cryoprotectants containing glycerol were associated with higher cumulus cell viability. However, the glycerol-egg yolk combination yielded the highest cell viability.
CONCLUSIONS: The cumulus comet assay demonstrated similar DNA integrity in cells frozen in each of the three cryoprotectants. The glycerol-egg yolk medium had the highest cell viability with little or no DNA damage after freeze-thaw. More studies are needed to examine the long-term effect of the cryoprotectants on thawed cumulus cell viability.
Authors:
E M Lindley; J D Jacobson; J Corselli; A King; P J Chan
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Journal of assisted reproduction and genetics     Volume:  18     ISSN:  1058-0468     ISO Abbreviation:  J. Assist. Reprod. Genet.     Publication Date:  2001 Oct 
Date Detail:
Created Date:  2001-11-08     Completed Date:  2002-04-12     Revised Date:  2013-06-09    
Medline Journal Info:
Nlm Unique ID:  9206495     Medline TA:  J Assist Reprod Genet     Country:  United States    
Other Details:
Languages:  eng     Pagination:  534-8     Citation Subset:  IM    
Affiliation:
Department of Gynecology and Obstetrics, Loma Linda University School of Medicine, 11370 Anderson Street Suite 3950, Loma Linda, California 92354, USA.
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MeSH Terms
Descriptor/Qualifier:
Acridine Orange / chemistry
Cells, Cultured
Coculture Techniques / methods*
Comet Assay
Cryopreservation / methods*
Cryoprotective Agents / pharmacology
DNA Damage*
Female
Fertilization in Vitro*
Fluorescent Dyes / chemistry
Glycerol / pharmacology
Humans
Male
Ovarian Follicle / cytology*
Pregnancy
Chemical
Reg. No./Substance:
0/Cryoprotective Agents; 0/Fluorescent Dyes; 56-81-5/Glycerol; 65-61-2/Acridine Orange
Comments/Corrections

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