Practical Selection Methods for Rat and Mouse Round Spermatids Without DNA Staining by Flow Cytometric Cell Sorting
Practical Selection Methods for Rat and Mouse Round Spermatids Without DNA Staining by Flow Cytometric Cell Sorting
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DOI：
10.1002/mrd.22644
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发表时间：
2016-06-01
影响因子：
2.5
通讯作者：
Nakauchi, Hiromitsu
中科院分区：
文献类型：
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作者：
Hayama, Tomonari;Yamaguchi, Tomoyuki;Nakauchi, Hiromitsu
将圆形精子细胞注射（ROSI）到未受精的卵母细胞中，可以使患有严重精子发生障碍的男性生育孩子。该技术在临床应用的一个限制是从睾丸组织中识别和分离圆形精子细胞。在这里，我们开发了一种有效且简单的方法，根据 DNA 含量（用 Hoechst 33342 或 Dye Cycle Violet 染色）或细胞直径和粒度（前向和侧向散射），使用流式细胞仪细胞分选来分离啮齿动物单倍体圆形精子细胞。 ROSI是用流式细胞术选择的圆形精子细胞进行的，我们从未染色的细胞中获得了健康的后代。因此，这种非侵入性方法可能成为饲养家畜和治疗人类男性不育症的有效选择。
Round spermatid injection (ROSI) into unfertilized oocytes enables a male with a severe spermatogenesis disorder to have children. One limitation of the application of this technique in the clinic is the identification and isolation of round spermatids from testis tissue. Here we developed an efficient and simple method to isolate rodent haploid round spermatids using flow cytometric cell sorting, based on DNA content (stained with Hoechst 33342 or Dye Cycle Violet) or by cell diameter and granularity (forward and side scatter). ROSI was performed with round spermatids selected by flow cytometry, and we obtained healthy offspring from unstained cells. This non- invasive method could therefore be an effective option for breeding domestic animals and human male infertility treatment.