Back to Search
Start Over
Development of a cryopreservation protocol for long-term storage of black tiger shrimp (Penaeus monodon) spermatophores.
- Source :
-
Theriogenology [Theriogenology] 2007 Nov; Vol. 68 (8), pp. 1192-9. Date of Electronic Publication: 2007 Sep 27. - Publication Year :
- 2007
-
Abstract
- The objectives of this study were to determine the effect of cryoprotectants on sperm viability and develop a freezing protocol for long-term storage of P. monodon spermatophores. Spermatophores suspended for 30 min in calcium-free saline (Ca-F saline) containing the cryoprotectants dimethyl sulfoxide (DMSO), ethylene glycol (EG), 1,2-propylene glycol (PG), formamide, and methanol at concentrations of 5, 10, 15, or 20% were studied using a modified eosin-nigrosin staining technique. The smallest reductions in apparent sperm viability occurred with DMSO; therefore, a freezing protocol was developed using Ca-F saline containing 5% DMSO. Spermatophores were cryopreserved using three protocols; cooling to a final temperature of -30, -80 or -80 degrees C and immediately stored in liquid nitrogen (cooling rates of -2, -4, -6, -8, -10, -12, -14 or -16 degrees C/min). Frozen spermatophores were thawed (2 min) at 30, 60, 70, or 90 degrees C. Successful cryopreservation of spermatophores in liquid nitrogen was achieved by a one-step cooling rate of -2 degrees C/min between 25 and -80 degrees C before storing in liquid nitrogen. Optimal thawing was in a 30 degrees C water bath for 2 min; this yielded live sperm after storage in liquid nitrogen for 210 days. Average sperm viability for fresh (97.8+/-2.9%) and cryopreserved spermatophores held for less than 60 days (87.3+/-4.1%) did not differ (P>0.05); however, that for spermatophores stored in liquid nitrogen between 90 and 210 days were lower (P<0.05) and varied from 27.3+/-3.4 to 53.3+/-4.3%. Thawed spermatophores previously held in liquid nitrogen for less than 62 days fertilized eggs (fertilization and hatching rates of 71.6-72.2% and 63.6-64.1%, respectively) at rates comparable to fresh spermatophores (70.8-78.2% and 66.3-67.8%, respectively). In conclusion, sperm within cryopreserved spermatophores stored in liquid nitrogen retained their viability for up to 210 days.
- Subjects :
- Animals
Dimethyl Sulfoxide toxicity
Ethylene Glycol toxicity
Fertilization in Vitro drug effects
Fertilization in Vitro veterinary
Formamides toxicity
Male
Methanol toxicity
Propylene Glycol toxicity
Semen Preservation methods
Sodium Chloride
Time Factors
Cryopreservation veterinary
Cryoprotective Agents toxicity
Penaeidae physiology
Semen Preservation veterinary
Spermatogonia drug effects
Subjects
Details
- Language :
- English
- ISSN :
- 0093-691X
- Volume :
- 68
- Issue :
- 8
- Database :
- MEDLINE
- Journal :
- Theriogenology
- Publication Type :
- Academic Journal
- Accession number :
- 17900683
- Full Text :
- https://doi.org/10.1016/j.theriogenology.2007.08.024