Journal of Assisted Reproduction and Genetics

, Volume 31, Issue 8, pp 983–991 | Cite as

Development of a high-yield technique to isolate spermatogonial stem cells from porcine testes

  • Min Hee Park
  • Ji Eun Park
  • Min Seong Kim
  • Kwon Young Lee
  • Hye Jin Park
  • Jung Im Yun
  • Jung Hoon Choi
  • Eun song Lee
  • Seung Tae Lee
Technological Innovations



To date, the methods available for isolating spermatogonial stem cells (SSCs) from porcine testicular cells have a low efficiency of cell separating. Therefore, we tried to develop a novel isolation technique with a high-yield cell separating ability to isolate SSCs from porcine testes.


We confirmed the presence of SSCs by measuring alkaline phosphatase (AP) activity and SSC-specific gene expression in neonatal porcine testis-derived testicular cells. Subsequently, the isolation of SSCs from testicular cells was performed using different techniques as follows: differential plating (DP), double DP, Petri dish plating post-DP, magnetic-activated cell sorting (MACS), and MACS post-DP. Positive AP staining was used to assess and compare the isolation efficiency of each method.


Petri dish plating post-DP resulted in the highest isolation efficiency. The putative SSCs isolated using this method was then further characterized by analyzing the expression of SSC-specific genes and -related proteins, and germ cell-specific genes. OCT4, NANOG, EPCAM, THY1, and UCHL1 were expressed transcriptionally, and OCT4, NANOG, SOX2, TRA-1-60, TRA-1-81, and PLZF were expressed translationally in 86 % of the isolated SSCs. In contrast, no difference was observed in the percentage of cells expressing luteinizing hormone receptor (LHR), a Leydig cell-specific protein, or GATA4, a Sertoli cell-specific protein, between SSCs and negative control cells. In addition, transcriptional expression of VASA, a primordial germ cell-specific marker, and DAZL, a premeiotic germ cell-specific marker, wasn’t and was detected, respectively.


We successfully developed a novel high-yield technique to isolate SSCs from porcine testes to facilitate future porcine SSC-related research.


Porcine Spermatogonial stem cells Testis Cell sorting 



This research was supported by Agricultural Biotechnology Development Program (IPET112015-4), Ministry of Agriculture, Food and Rural Affairs, Republic of Korea.


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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Min Hee Park
    • 1
  • Ji Eun Park
    • 2
  • Min Seong Kim
    • 1
  • Kwon Young Lee
    • 3
  • Hye Jin Park
    • 1
  • Jung Im Yun
    • 3
  • Jung Hoon Choi
    • 3
  • Eun song Lee
    • 3
  • Seung Tae Lee
    • 1
    • 2
    • 4
  1. 1.Department of Animal Life ScienceKangwon National UniversityChuncheonSouth Korea
  2. 2.Division of Applied Animal ScienceKangwon National UniversityChuncheonSouth Korea
  3. 3.College of Veterinary Medicine and Institute of Veterinary ScienceKangwon National UniversityChuncheonSouth Korea
  4. 4.Department of Animal Life Science and Division of Applied Animal ScienceKangwon National UniversityChuncheonSouth Korea

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