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- Title
Flow Cytometric Sorting of Fresh and Frozen-Thawed Spermatozoa in the Western Lowland Gorilla (Gorilla gorilla gorilla).
- Authors
O'Brien, J. K.; Stojanov, T.; Crichton, E. G.; Evans, K. M.; Leigh, D.; Maxwell, W. M. C.; Evans, G.; Loskutoff, N. M.
- Abstract
We adapted flow cytometry technology for high-purity sorting of X chromosome-bearing spermatozoa in the western lowland gorilla (Gorilla gorilla gorilla). Our objectives were to develop methodologies for liquid storage of semen prior to sorting, sorting of liquid-stored and frozen-thawed spermatozoa, and assessment of sorting accuracy. In study 1, the in vitro sperm characteristics of gorilla ejaculates from one male were unchanged (P>0.05) after 8 hr of liquid storage at 15°C in a non-egg yolk diluent (HEPES-buffered modified Tyrode's medium). In study 2, we examined the efficacy of sorting fresh and frozen-thawed spermatozoa using human spermatozoa as a model for gorilla spermatozoa. Ejaculates from one male were split into fresh and frozen aliquots. X-enriched samples derived from both fresh and frozen-thawed human semen were of high purity, as determined by fluorescence in situ hybridization (FISH; 90.7%±2.3%, overall), and contained a high proportion of morphologically normal spermatozoa (86.0%±1.0%, overall). In study 3, we processed liquid-stored semen from two gorillas for sorting using a modification of methods for human spermatozoa. The sort rate for enrichment of X-bearing spermatozoa was 7.3±2.5 spermatozoa per second. The X-enriched samples were of high purity (single-sperm PCR: 83.7%) and normal morphology (79.0%±3.9%). In study 4 we examined frozen-thawed gorilla semen, and the sort rate (8.3±2.9 X-bearing sperm/sec), purity (89.7%), and normal morphology (81.4%±3.4%) were comparable to those of liquid-stored semen. Depending on the male and the type of sample used (fresh or frozen-thawed), 0.8–2.2% of gorilla spermatozoa in the processed ejaculate were present in the X-enriched sample. These results demonstrate that fresh or frozen-thawed gorilla spermatozoa can be flow cytometrically sorted into samples enriched for X-bearing spermatozoa. Am. J. Primatol. 66:297–315, 2005. © 2005 Wiley-Liss, Inc.
- Subjects
SPERMATOZOA; WESTERN lowland gorilla; EJACULATION; ANIMAL sexual behavior; ANIMAL behavior; FLUORESCENCE in situ hybridization
- Publication
American Journal of Primatology, 2005, Vol 66, Issue 4, p297
- ISSN
0275-2565
- Publication type
Article
- DOI
10.1002/ajp.20158