Asghar Talebian
- Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Mohammad Reza Sadeghi
- Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Hooman Sadri-Ardekani
- Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Masoumeh Bolourzadeh
- Avicenna Infertility Clinic, Avicenna Research Institute (ACECR), Tehran, Iran
Mohammad Mehdi Akhondi Corresponding Author
- Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran

Received: 4/1/2005 Accepted: 4/1/2005 - Publisher : Avicenna Research Institute

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Introduction: Cryopreservation is a branch of cryo biology concern with the maintenance of cells during prolonged storage in the frozen state at ultra low temperature. Sperm cryopreservation is routinely perfor-med in andrology laboratory and fertility centers and industry.Cryopreservation decrease sperm quality and its Fertilizing capacity. Evaluation of different sperm cryo-media and methods have been performed for several years. Yet, it is not known the best media and technique for sperm cryopreservstion. Thus it seems very necessary to set a sperm cryo-media and method base on experimental finding. Therefore, the aim of this study is the assessment of sperm survival and motility during cryopreservation by three different media including HSPM (Human Sperm Preservation Medium), TYBG (Test-Yolk Buffer) and GEYC (Glycerol Egg Yolk Citrate) and two different cryo-techniques including programmable (with apparatus) and using vapor phase (without apparatus). Materials and Methods: 22 samples were collected of normal male into sterile container. Sperm analysis was performed on the base of WHO criteria and freezing program including programmable and vapor pha-se was performed. Prefreezing sperm concentration and motility was evaluated before and after freezing and subsequently survival rate (CSF) was calculated. Each sample was divided in three parts and mixed with three different medium, subsequently at least two straws filled of each sample. One straw freezed with equipment and another one in vapor phase. The SPSS (Edition 11.0) statistical program was utilized for statistical analysis and t-test was used to compare sperm motility prefreezing and post thawing with two different cryopreservation methods and ANOVA/LSD was used to compare sperm motility with three different cryopreservation media. Significant level was considered p<0.05. results: base on our result, sperm motility was %46.138.29 prefreezing , but considerably decreased post thawing by programmable technique in hspm (%16.95), geyc (%16.314.57) and tybg (%16.04 4.75) and also by using vapor phase technique in hspm (%16.954.55), geyc(%14.135.14) and tybg (%14.184.47). there was significantly difference between programmable and vapor phase technique in geyc (p="0.001)" and tybg (p="0.007)." there was also significant difference between hspm and both other media (p="0.05)." conclusion: according to our study, sperm thawing after its freezing by liquid nitrogen vapor phase in three different media (hspm, geyc, tybg) leads to significant reduction in sperm motility. the best result in sperm motility and survival achieved through programmable thawing with apparatus and in comp-aring freezing media, hspm media significantly has higher survival rate in comparison with tybg and geyc media (p-value="0.05)." since the freezing apparatus is not available in most centers therefore, utilizing hspm freezing media and vapor phase freezing method would be recommended for normal semen samples.< pan>

Keywords: Sperm bank, Cryopreservation, Cryo-media, HSPM, GEYC, TYBG, Slow freezing, Vapor phase

To cite this article:


  1. Mortimer D. Practical Laboratory Andrology. Oxford University Press incorporated. Chapt. 14, Semen cryo-preservation.1994;pp:301-323.
  2. Shaw J.M., Oranratnachai A., Trounson A.O. Cryopre-servation of oocytes and embryos. Chapter 16. In: Handbook of Invitro Fertilization Second Edition. Edit-ed by Alan O. Trounson and David K. Gardner. CRC press Inc.2000.
  3. Brotherton J. Cryopreservation of human semen. Arch of Androl.1990;25:181-195.
  4. Mahony M.C., Morshedi M., Scott R.T., De Villiers A. Erasmus E. Role of spermatozoa cryopreservation in assisted reproduction. In: human spermatozoa in assis-ted reproduction. Edited by Acosta A.A., Swanson R. J., Ackerman S.B., kruger T.A.F., Van Zyl J.A., and Menkveld R. Baltimore: Williams & wilkins Co., Cha-pt. 10, pp:100,1990.
  5. Anger J.T., Gilbert B.R. Goldstein M. Cryopreservation of sperm: Indications, Methods and Results. J Urol. 2003;170:1079-1084.
  6. Jahnel F. Uber die wiederstandsfahigkeit von mens-chlichen spermatozoan gegenuber starke kalte. Klin Wschr.1937;17:1273-1275.
  7. Polge G., Smith A.U., Parkes A.S. Retrival of (bovine) spermatozoa after vitrification and dehydration at low temperatures. Nature.1949; 164:666-669.
  8. Bunge R.G., Sherman J.K. Fertilizing capacity of fro-zen human spermatozoa. Nature.1953;172: 767.
  9. Prins G.S., Weidel L. A comparative study of buffer systems as cryoprotectants for human spermatozoa. Fertil Steril.1986;46(1):147-149.
  10. Mahadevan M., Trounson A.O. Effect of cryoprotec-tive media and dilution methods on the preservation of human spermatozoa. Andrologia.1983;15(4):355-366.
  11. Stanic P., Tandara M., Sonicki Z., Simunic V., Radak-ovic B., Suchanek E. Comparison of protective media and freezing techniques for cryopreservation of human semen. Eur J Obstet Gynecol Reprod Biol.2000;91:65-70.
  12. Nallella K.P., Sharma R.K., Allamaneni S.S.R., Aziz N., Agarwal A. Cryopreservation of human spermato-zoa: comparison of two cryopreservation methods and there cryoprotectants. Fertil Steril.2004;82(4):913-918.
  13. Gardner D.K., Weissman A., Howles C.M., Shoham Z. Assisted Reproductive Techniques. Chapt. 5: Evalua-tion of sperm.2001;P:61-76.
  14. Hammadeh M.E., Georg T., Rosenbaum P., Schmidt W. Association between freezing agent and acrosome damage of human sepermatozoa from subnormal and normal semen. Andrologia. 2001;33:331-336.
  15. Baratz S.L., Rothschild E., Grach B., Koifman M., Shiloh H., Ishai D. Dirnfeld M. The value of sperm pooling and cryopresenvation in patients with transi-ent azoospermia or sever oligoasthenoteratozoosper-mia. Hum Reprod.2002;17(1):157-160.
  16. McLaughlin E.A., Ford W.C.L., Hull M.G.R. A com-parison of the freezing of human semen in the uncircu-lated vapour above liquid nitrogen and in a comer-cial semi programmable freezer. Hum Reprod.1990;5:724-8.
  17. Ragni G., Caccamo A.M., Dalla Serra A., Guercilena S. Computerized slow staged freezing of semen from men with testicular tumors or Hodgkin’s disease prese-rves sperm better than standard vapor freezing. Fertil Steril.1990;53: 1072-5.
  18. Thachill J.V., Jewett M.A.S. Preservation techniques for human semen. Fertil Steril.1981; 35:546-8.
  19. Critser J.K., Kuse Benda A.R., Aaker D.V., Arneson B.W., Ball G.D. Cryopreservation of human spermato-zoa III. The effect of cryoprotectants on motility. Fertil Steril.1988;50:314-20.
  20. Serafini P., Mars R.P. Computerized stage-freezing technique improves sperm survival and preserve pene-tration of zona-free hamster ova. Fertil Steril.1986;45: 854-858.
  21. Critser J.K., Arenson B.W., Aaker D.V., Huse-Benda A.R., Ball G.D. Cryopreservation of human spermatozoa II. Postthaw chronology of motility and of zona-free hamster ova penetration. Fertil Steril.1987;47:980-984.
  22. Tulandi T., McInnes R.A. Vaginal Lubricant: effect of glycerol and egg white on human sperm motility and progression in vitro. Fertil Steril 1984;41:151-153.
  23. Weidel L., Prins G.S. Cryopreservation of human spermatozoa frozen in eight different buffer system. J Androl.1987;8:41-47.
  24. Watson P.F. The preservation of semen in mammals. In: Oxford reviews of reproductive biology. Finn CA (Eds) Oxford University Press, Oxford.1979;pp:283-350.
  25. Gao D.Y., Ashworth E., Watson P.F. Hypersmotic tole-rance of human spermatozoa: separate effects of glyce-rol, sodium chloride and sucrose on spermiolysis. Biol Reprod.1993;49:112-113.
  26. Jezek D., Schulze W., Kalanj-Bognar S., Vukelic Z., Milavec-Puretic V., krhen I. Effects of various cryo preservation media and freezing thawing on the morphology of rat testicular biopsies. Andrologia.2001; 33:368-378.
  27. Sawada Y. preservation of human semen by deep freezing. Int J Fert.1964;9:525-532.
  28. Sawada Y., Ackerman D.R. Use of frozen semen. In: S.J. Behrman and R.W. kistner (Editors) progress in infertility. Boston: little Brown.1968;pp:731-748.
  29. Jeyendran R.S., Graham E.F. An evaluation of cryo-protective compounds on bovine spermatozoa. Cryo-biology.1980;17:458-464.
  30. Polge C. Freezing of spermatozoa. In: Ashwood Smith and J. Farrant (Eds): Low temperature presser-vation in medicine and biology. Bath: Pitman.1980; pp:45-64.
  31. Richardson D.W. Techniques of sperm storage. In: Artificial insemination. R.C.O.G., London.1976;pp:97-125.
  32. Mann T. Biochemistry of semen and of the male reproductive tract. London. Methuen. (2nd Edition).1964.
  33. Graham E.F., Crabo B.E., Brown K.L. Effect of some zwitterions buffers on the freezing and storage of sper-matozoa. J Dairy Sci.1972;55:372-8.
  34. Mc Gonagle L.S., Goldstein M., Feld schuh J. Foote R.H. The influence of cryprotective media and processing procedures on motility and migration of frozen thawed human sperm. Asian J Androl.2002; 4:137-141.
  35. Sherman J.K. Cryopreservation of human semen. In: Keel B.A., Webster B.W. (Editors); CRC Handbook of the laboratory Diagnosis and Treatment of infertility CRC press. Boca Raton, FL.1990.
  36. Emmens. C.W., Blackshaw A.W. Artificial insemina-tion. Physiol Rev.1971;36: 227-306.
  37. Luyet B., Masat R. On the cryoprotective action of albumin on erythocytes. Cryobiology. 1967;3:370.
  38. Glaub J.C., Milis R.N., Katz D.F. Improved motility recovery of human spermatozoa after freeze preserva-tion via a new approach. Fertil Steril.1976;27:1283-1291.
  39. Katayama K.S., Stehlik E., Roesler M., Jeyedran R.S., Holmgren W.J., Zaneveld L.J. Treatment of human spermatozoa with an egg yolk medium can enhance the outcome of in vitro fertilization. Fertil Steril.1989; 52:1077-1079.
  40. Barak Y., Amit A., Lessing J., Paz G., Homonnai Z., Yogev L. Improved fertilization rate in an in vitro fertilization program by egg yolk-treated sperm. Fertil Steril.1992;58:197-198.
  41. McCoshen J.A., Wodzicki A., Tyson J.E. Effectiveness of human semen frozen in TESE-yolk- buffered medium on AID outcome. Fertil Steril.1984;42:164.
  42. Griffin J.E.T., Nunn W.R., Hartigen P.J. An immune response to egg yolk semen diluent in dairy cows. J Reprod Fert.1971;25:193-199.
  43. Park Y.W, Hunter A.G. Effect of repeated inseminations with egg yolk semen extender on fertility in cattle. J. Dairy. Sci.1977;60:1645-1649.
  44. Jondet M. Congelation du sperme humain. Bull Acad Vet.1976;49:373-384.
  45. Foote R.H. Fertility of bull semen at high extension rates in Tris buffered extenders. In: Reproduzione animale e fecondazione artificiale. Bologna: Edizione Agricole. 1972;pp:99-105.
  46. Van den Berg L., Rose D. Effect of freezing on the pH and composition of sodium and potassium phosph-ate solution: the reciprocal system KH2PO4- Na2HPO4-H2O. Arch Biochem Biophys.1959;81:319-329.


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