use of the acridine orange test and the TUNEL assay to assess the integrity of freeze-dried bovine spermatozoa DNA

The ability to detect nuclear damage is an important tool for the development of sperm preservation methods. We used the acridine orange test (AOT) and the terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling (TUNEL) assay to assess the DNA status of sperm cells preserved with diffe...

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Published inGenetics and molecular research Vol. 6; no. 1; pp. 94 - 104
Main Authors Martins, C.F, Dode, M.N, Báo, S.N, Rumpf, R
Format Journal Article
LanguageEnglish
Published Brazil 01.01.2007
Subjects
acridine orange
animal breeding
Animals
Cattle
Chromatin - chemistry
Chromatin - metabolism
Coloring Agents
DNA - chemistry
DNA - metabolism
DNA damage
DNA Fragmentation
fertilization (reproduction)
Flow Cytometry
Freeze Drying
In Situ Nick-End Labeling
Male
Nellore
Nucleic Acid Conformation
Protein Conformation
Semen Preservation - adverse effects
Semen Preservation - methods
spermatozoa
Spermatozoa - chemistry
Staining and Labeling
terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling assay
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Summary:The ability to detect nuclear damage is an important tool for the development of sperm preservation methods. We used the acridine orange test (AOT) and the terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling (TUNEL) assay to assess the DNA status of sperm cells preserved with different lyophilization media. The AOT did not detect any differences between different lyophilization media. However, differences in DNA integrity were observed among treatments with the TUNEL assay, suggesting that TUNEL is a more sensitive method to evaluate sperm DNA. The use of TCM 199 and 10% FCS as a lyophilization medium resulted in 14% of the cells with DNA fragmentation in TUNEL test. The AOT indicated only 4% of the cells with chromatin damage, with this same treatment, with no significant differences when compared to the other treatments. The degree of DNA fragmentation was negatively related to fertilizing potential, as sperm DNA damage was inversely correlated with pro-nucleus formation. The TUNEL assay was found to be an efficient method to detect DNA damage in sperm, and it could be used as a tool to predict male fertility.
Bibliography:http://funpecrp.com.br/gmr/index.htm
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ISSN:1676-5680
1676-5680