title: The effect of repeated freezing and thawing on human sperm DNA fragmentation.
abstract: OBJECTIVE
  To investigate the effects of repeated freezing and thawing on human sperm motility, vitality, and DNA integrity.
  
  
  METHODS
  A prospective clinical study.
  
  
  METHODS
  Tertiary care fertility clinic.
  
  
  METHODS
  Twenty men presenting for infertility investigations.
  
  
  METHODS
  Each sample was subjected to three cycles of freezing and thawing both with and without washing steps and the addition of fresh cryoprotectant between each cycle.
  
  
  METHODS
  Percentage sperm DNA fragmentation, motility, and vitality before and following repeated freezing and thawing.
  
  
  RESULTS
  The percentage sperm DNA fragmentation rose significantly following each freeze-thaw cycle; however, samples that were not washed and to which fresh cryoprotectant was not added after each cycle fared significantly better than their washed counterparts. Both motility and vitality decreased steadily following each cycle but cell survival was significantly greater in the unwashed samples.
  
  
  CONCLUSIONS
  In terms of the level of sperm DNA fragmentation, up to three cycles of freezing and thawing can be performed with a level of risk comparable to that following a single cycle of freezing and thawing. This is provided that samples are refrozen in their original cryoprotectant and not washed or altered in any way in between, and provided that they are separated by density gradient centrifugation or swim-up before use in assisted reproduction technology.
authors.affiliation: Fertility First, P.O. Box 807, Hurstville NSW 2220, Australia. lthomson@fertilityfirst.com.au
authors.forename: Laura Kelly
authors.initials: LK
authors.lastname: Thomson
authors.forename: Steven Denis
authors.initials: SD
authors.lastname: Fleming
authors.forename: Katrina
authors.initials: K
authors.lastname: Barone
authors.forename: Julie-Anne
authors.initials: JA
authors.lastname: Zieschang
authors.forename: Anne Melton
authors.initials: AM
authors.lastname: Clark
mesh_terms: D000328:Adult; D002499:Centrifugation, Density Gradient / Q000379:methods / Q000592:standards; D015925:Cryopreservation / Q000379:methods* / Q000592:standards; D003451:Cryoprotective Agents / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D005615:Freezing; D006801:Humans; D007248:Infertility, Male / Q000503:physiopathology / Q000628:therapy; D008297:Male; D008875:Middle Aged; D011446:Prospective Studies; D012662:Semen Preservation / Q000009:adverse effects / Q000379:methods* / Q000592:standards; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology*
chemical_list: D003451:Cryoprotective Agents