Fertility Center of CHA Gangnam Medical Center, College of Medicine, CHA University, Seoul, South Korea.
Fertil Steril. 2013 Dec;100(6):1564-71.e1-5. doi: 10.1016/j.fertnstert.2013.08.017. Epub 2013 Sep 11.
To determine whether ligation-mediated real-time polymerase chain reaction (LM-RT-PCR), which combines LM-PCR, and RT-PCR, can detect sperm DNA fragmentation (DF) in human semen samples.
Three-way comparison of terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), sperm chromatin dispersion (SCD), and LM-RT-PCR for detecting sperm DNA fragmentation.
University hospital-based research laboratory.
PATIENT(S): Twenty-five men presenting at an infertility clinic.
INTERVENTION(S): Basic analysis of sperm concentration, motility, vitality, and morphology, with each semen sample equally divided into three aliquots that were evaluated for fragmentation using TUNEL, SCD, and LM-RT-PCR assays.
MAIN OUTCOME MEASURE(S): In TUNEL and SCD assays, counts of the number of sperm with tetramethylrhodamine (TMR) red signals or no halo; in LM-RT-PCR results, evaluation of the threshold cycles (Ct) and relative fluorescence unit (RFU) values.
RESULT(S): The median percentage of sperm with positive results for fragmentation in the TUNEL and SCD assays were 20.5% and 20.7%, respectively. To compare the accuracy of the TUNEL, SCD, and LM-RT-PCR assays, we divided the semen samples into two groups according to the TUNEL results: low and high percentage of sperm fragmentation. In the LM-RT-PCR results, the values of the cycles of threshold (Ct) and relative fluorescence unit (RFU) statistically significantly differed between the low and high percentage of sperm fragmentation groups. Comparisons among the TUNEL, SCD, and LM-RT-PCR assays revealed that the correlation patterns according to DNA fragmentation were similar in both the groups with high and low percentage of DNA fragmentation. Our morphologic analysis indicated that the fragmentation of sperm DNA did not appear to influence sperm morphology.
CONCLUSION(S): These results indicate that the LM-RT-PCR technique is another useful tool for detecting DNA fragmentation, a parameter of sperm quality in human semen alone or combined with TUNEL or SCD assays.
确定连接介导的实时聚合酶链反应（LM-RT-PCR）是否可以检测人类精液样本中的精子 DNA 碎片化（DF）。
三向比较末端脱氧核苷酸转移酶 dUTP 缺口末端标记（TUNEL）、精子染色质弥散（SCD）和 LM-RT-PCR 检测精子 DNA 碎片化。
以大学医院为基础的研究实验室。
25 名在不孕不育诊所就诊的男性。
基本分析精子浓度、活力、活力和形态，每个精液样本均等分为三份，分别用 TUNEL、SCD 和 LM-RT-PCR 评估碎片。
在 TUNEL 和 SCD 试验中，四甲基罗丹明（TMR）红色信号或无晕的精子计数；在 LM-RT-PCR 结果中，评估阈值循环（Ct）和相对荧光单位（RFU）值。
TUNEL 和 SCD 试验中精子碎片阳性结果的中位数百分比分别为 20.5%和 20.7%。为了比较 TUNEL、SCD 和 LM-RT-PCR 试验的准确性，我们根据 TUNEL 结果将精液样本分为两组：精子碎片化的低百分比和高百分比。在 LM-RT-PCR 结果中，低和高百分比精子碎片化组之间的阈值循环（Ct）和相对荧光单位（RFU）值的差异具有统计学意义。TUNEL、SCD 和 LM-RT-PCR 试验之间的比较表明，根据 DNA 碎片化的相关性模式在高和低 DNA 碎片化百分比的组中相似。我们的形态学分析表明，精子 DNA 的碎片化似乎不会影响精子形态。
这些结果表明，LM-RT-PCR 技术是另一种有用的工具，可单独或与 TUNEL 或 SCD 检测联合检测人类精液中精子质量的 DNA 碎片化参数。
