Isolation, Cryopreservation, and Transplantation of Spermatogonial Stem Cells

  • Nilam Sinha
  • Eoin C. WhelanEmail author
  • Ralph L. Brinster
Part of the Methods in Molecular Biology book series (MIMB, volume 2005)


Spermatogonial stem cell (SSC) culture and transplantation pave the way for clinical restoration of fertility in male prepubertal cancer survivors. In this chapter we detail the steps for isolating and freezing testicular tissue along with protocols for the subsequent recovery from cryopreservation and transplantation of cells into a recipient testis. Transplantation of cultured or thawed SSCs provides not only a functional assay for identification of stem cells, a critical tool for the study of the germline stem cell niche in model organisms, but also a framework for reconstitution of spermatogenesis in humans. As proof of concept, the outlined methods have been performed successfully in the murine model and have the potential to be translated to clinical environments.

Key words

Spermatogonial stem cells Germ cell Testicular biopsy Cryopreservation Transplantation Microinjection Spermatogenic colony Infertility Prepubertal cancer 



We thank James Hayden, RBP, FBCA, for assistance in preparation of figures and Mary Avarbock for advice and expert assistance. This work was supported by the National Institute of Child Health and Human Development (R.L.B.) and the Robert J. Kleberg, Jr. and Helen C. Kleberg Foundation (R.L.B.).


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Nilam Sinha
    • 1
  • Eoin C. Whelan
    • 1
    Email author
  • Ralph L. Brinster
    • 1
  1. 1.Department of Biomedical SciencesSchool of Veterinary Medicine, University of PennsylvaniaPhiladelphiaUSA

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