title: Isolation of germ cells from human testicular tissue for low temperature storage and autotransplantation.
abstract: OBJECTIVE
  To develop a new protocol for conserving fertile potential in men undergoing sterilizing chemotherapy by low temperature banking of germ cells which can be returned to the patient's testes after thawing.
  
  
  METHODS
  Isolation of human and murine germ cells for comparing cellular viability after cooling to liquid nitrogen temperatures by the use of different cryoprotective agents and for infusion into the testis.
  
  
  METHODS
  Laboratory research environment.
  
  
  METHODS
  Men undergoing routine surgery in a urology department.
  
  
  METHODS
  Testicular biopsy.
  
  
  METHODS
  Cellular viability and infusion of seminiferous tubules.
  
  
  RESULTS
  After isolation using a two-step enzymatic disaggregation protocol, 66% to 87% of germ cells from human and murine specimens, respectively, were still viable. Cell survival was similar in four commonly used cryoprotective agents after cooling to liquid nitrogen temperatures. Seminiferous tubules infused by back flow with dye solution via the rete testis were filled with an efficiency of 55%.
  
  
  CONCLUSIONS
  Judging from the high viability of unfractionated germ cells, it is feasible to isolate germ cells from testicular biopsies for low temperature banking with the aim of attempting to restore fertility after iatrogenic sterilization.
authors.affiliation: Centre for Reproduction, Growth, and Development, Leeds General Infirmary, University of Leeds, United Kingdom.
authors.forename: P F
authors.initials: PF
authors.lastname: Brook
authors.forename: J A
authors.initials: JA
authors.lastname: Radford
authors.forename: S M
authors.initials: SM
authors.lastname: Shalet
authors.forename: A D
authors.initials: AD
authors.lastname: Joyce
authors.forename: R G
authors.initials: RG
authors.lastname: Gosden
mesh_terms: D000818:Animals; D002469:Cell Separation; D017690:Cell Transplantation; D015925:Cryopreservation; D003451:Cryoprotective Agents; D006801:Humans; D007248:Infertility, Male / Q000139:chemically induced / Q000601:surgery; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D012671:Seminiferous Tubules; D013094:Spermatozoa / Q000166:cytology*; D013737:Testis / Q000166:cytology*
chemical_list: D003451:Cryoprotective Agents