Visualization and tracking of live type a spermatogonia using a fluorescence-conjugated antibody in Salmo species
Visualization and tracking of live type a spermatogonia using a fluorescence-conjugated antibody in Salmo species
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使用荧光偶联抗体对 Salmo 物种的活 A 型精原细胞进行可视化和跟踪
DOI：
10.1016/j.aquaculture.2020.736096
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发表时间：
2021
期刊：
影响因子：
4.5
通讯作者：
Yoshizaki Goro
中科院分区：
文献类型：
--
作者：
Ichida Kensuke;Matsushita Yoshiyuki;Amano Yuichi;Miwa Misako;Nagasawa Kazue;Hayashi Makoto;Mizutani Hanaka;Takahashi Momo;Boonanuntanasarn Surintorn;Yoshizaki Goro
将生殖细胞冷冻保存与移植相结合是保存各种鱼类遗传资源的有力工具。在该生殖细胞移植系统中，由于在各种睾丸细胞中只有A型精原细胞（ASG）具有定殖受体性腺和恢复配子发生的潜能，因此用于ASG的可视化、分离和追踪的技术便于操作。在这里，我们建立了ASG可视化技术在两个鲑鱼：褐鳟鱼和大西洋鲑鱼。我们应用单克隆抗体（抗体No.95），其特异性识别从虹鳟鱼获得的ASG的细胞表面抗原，以对抗Salmospecies。免疫组织化学研究表明，95号抗体还特异性地检测到褐鳟鱼和大西洋鲑鱼的ASG。此外，标记基因分析的褐鳟鱼睾丸细胞分选的流式细胞仪使用Alexa面粉488-结合的抗体95号显示，荧光阳性细胞具有ASGs的分子特征。最后，为了研究抗体阳性细胞的可移植性，将用95号抗体预染色的ASG移植到虹鳟鱼幼体的腹腔中。结果，荧光细胞掺入受体性腺中，表明可视化的ASG具有可移植性。因此，本研究中开发的ASG可视化方法将有助于开发更有效的鲑鱼生殖细胞移植技术。
Combining germ cell cryopreservation with transplantation is a powerful tool for preserving the genetic resources of various fish strains. In this germ cell transplantation system, since only type A spermatogonia (ASGs) among the various testicular cells have the potency to colonize recipient gonads and resume gametogenesis, techniques for visualization, isolation, and tracing of ASGs facilitate ease of manipulation. Here we established an ASG visualization technique in twoSalmospecies: brown trout and Atlantic salmon. We applied a monoclonal antibody (antibody No. 95), which specifically recognizes a cell surface antigen of ASGs obtained from rainbow trout, againstSalmospecies. Immunohistochemistry studies showed that antibody No. 95 also specifically detected the ASGs of brown trout and Atlantic salmon. Furthermore, marker-gene analyses of brown trout testicular cells sorted by flow cytometry using Alexa flour 488-conjugated antibody No. 95 revealed that fluorescent-positive cells possessed molecular characteristics of ASGs. Finally, to investigate the transplantability of antibody-positive cells, ASGs pre-stained with antibody No. 95 were transplanted into the peritoneal cavity of rainbow trout larvae. As a result, fluorescent cells incorporated into the recipient gonads, suggesting that the visualized ASGs possess transplantability. Thus, it is expected that the ASG visualization methods developed in this study will facilitate the development of more efficiency of germ cell transplantation techniques inSalmospecies.