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  • 11
    Electronic Resource
    Electronic Resource
    Flow sorting of X and Y chromosome-bearing spermatozoa into two populations (1987)
    New York, NY : Wiley-Blackwell
    Gamete Research 16 (1987), S. 1-9 
    Details
    ISSN: 0148-7280
    Keywords: flow cytometry ; DNA ; sperm separation ; fluorescent stain ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The only established difference on which to base the separation of X and Y chromosome-bearing spermatozoa is chromosomal constitution. This difference is quantifiable both from chromosome morphology (karyotype) and from DNA content. Flow cytometric techniques were used to measure relative DNA content of the X and Y populations and to flow-sort spermatozoa from Chinchilla laniger. Epididymal spermatozoa were recovered in PBS, fixed in 80% ethanol, treated with papain and dithioerythritol, and stained for DNA with Hoechst 33342. Sperm nuclei were analyzed and sorted on an EPICS V flow cytometer/cell sorter, modified specifically for spermatozoa. Two clearly resolved peaks (coefficient of variation 〈 1.5%) with approximately 7.5% difference in DNA content between X and Y chromosome-bearing spermatozoa were evident. Sperm nuclei were sorted from a portion of the X and Y peaks at a rate of 55 nuclei/sec for each population. Purities of individual X and Y populations averaged 95% as determined by reanalysis of the sorted populations. Successful sorting of Chinchilla X and Y chromosome-bearing spermatozoa into separate populations may aid in the identification of a biochemical marker that could be used to discriminate between the two sperm populations and lead to a practical procedure for sexing spermatozoa.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120160102
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  • 12
    Electronic Resource
    Electronic Resource
    The movement characteristics of rabbit spermatozoa before and after activation (1981)
    New York, NY : Wiley-Blackwell
    Gamete Research 4 (1981), S. 275-282 
    Details
    ISSN: 0148-7280
    Keywords: spermatozoa ; activation ; oviduct ; sperm movements ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Spermatozoa were flushed with mineral oil from the lower isthmus of the rabbit oviduct at four hours postcoitus (pc) and 11 hours pc. Videotapes were made of sperm behavior in the native isthmic fluid and after dilution of the fluid with culture medium. The tapes showed that, initially, spermatozoa in the native isthmic fluid were virtually immotile, but immediately resumed movement on contact with the culture medium. Isthmic sperm motility then evolved over a five- to 10-minute interval into the characteristic biphasic pattern of activated movement. Cine films of isthmic spermatozoa taken with a high-speed camera were analyzed to determine flagellar beat frequency, maximum flagellar curvature, and swimming velocity. Progressiveness ratios and hydrodynamic power outputs were then calculated for individual spermatozoa. Two phases of activated sperm movement, a whiplash phase and a progressive phase, were identified and characterized. The power output of activated spermatozoa increased twentyfold in comparison with the preactivated state. The power output of activated spermatozoa did not differ between the two phases of activated movement.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120040403
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  • 13
    Electronic Resource
    Electronic Resource
    Flow sorting of X and Y chromosome-bearing mammalian sperm: Activation and pronuclear development of sorted bull, boar, and ram sperm microinjected into hamster oocytes (1988)
    New York, NY : Wiley-Blackwell
    Gamete Research 21 (1988), S. 335-343 
    Details
    ISSN: 0148-7280
    Keywords: flow cytometry ; sperm separation ; DNA ; sex ratio ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Flow cytometric techniques were used to measure relative DNA content of X and Y chromosome-bearing bull, boar, and ram sperm populations and to separate the two sex-determining populations. Neat semen was prepared for flow cytometric analysis by washing, light sonication, and staining with 9 μM Hoechst 33342. Computer analysis of the bimodal histograms showed mean X-Y DNA differences of 3.9, 3.7, and 4.2% for bull, boar, and ram, respectively. Flow cytometric reanalysis of sorted bull, boar, and ram sperm showed purities greater than 90%. Bull, boar, and ram sperm nuclei were microinjected into hamster oocytes. Microinjected sperm were either unsorted, sorted, unsorted plus dithio-threitol (DTT) exposure, or sorted plus DTT exposure. Following microinjection, eggs were incubated 3 hr, fixed, and stained. A total of 579 eggs was observed for sperm activation (decondensation or formation of a male pronucleus). A lower percentage of sorted than unsorted (3 vs. 23%) boar sperm was activated (P 〈.05). However, sorted and unsorted DTT-exposed boar sperm or sorted and unsorted bull or ram sperm, regardless of DTT treatment, did not differ significantly. Sorted sperm nuclei of both rams and bulls exhibited higher activation rates than sorted boar sperm (P 〈.05). Treatment of sperm with DTT increased the activation rate (P 〈 .05) for sorted boar sperm but not for bull or ram sperm. These data represent the first separation of bull, boar, and ram X and Y chromosome-bearing sperm populations and the first evidence that sperm of domestic animals sorted on the basis of DNA by flow cytometric procedures have the ability to decondense and to form pronuclei upon injection into a hamster egg.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120210402
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  • 14
    Electronic Resource
    Electronic Resource
    Effect of liquid storage and cryopreservation of boar spermatozoa on acrosomal integrity and the penetration of zona-free hamster ova in vitro (1987)
    New York, NY : Wiley-Blackwell
    Gamete Research 16 (1987), S. 193-204 
    Details
    ISSN: 0148-7280
    Keywords: sperm penetration ; storage ; semen quality ; swine ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The effect of liquid storage and cryopreservation of boar spermatozoa on sperm motility, acrosomal integrity, and the penetration of zona-free hamster (ZFH) ova was examined. The sperm penetration assay (SPA) provides valuable information on specific events of fertilization and is a potentially useful indicator of sperm fertility. Ejaculated semen from 4 boars was subjected to 3 treatments: fresh (FRE, no storage), liquid-stored (LIS, stored at 18°C for 3 days), and frozen (FRO, frozen by pellet method and stored at -196°C for 3 days). A highly motile sperm population was isolated by the swim-up procedure (1 hr). FRE and LIS were incubated an additional 3 hr at 39°C in a Tris-buffered medium to elicit capacitation and the acrosome reaction. Sperm motility and acrosomal integrity were assessed before and after incubation. For the SPA, sperm and eggs were incubated at 39°C for 3 hr in Hams F-10 medium. Each egg was assessed for sperm penetration, sperm binding, and stage of development. Percentages of sperm motility and sperm with a normal apical ridge (NAR) prior to incubation were 78 and 78 (FRE), 75 and 69 (LIS), and 28 and 50 (FRO). After incubation, percentages of motility, NAR, and acrosome-reacted sperm were 34, 10, and 73 (FRE); 43, 24, and 51 (LIS); and 18, 13, and 59 (FRO). A somewhat higher (P 〈 .05) percentage of ZFH ova was penetrated by FRE (45.8) than by LIS (42.0). Penetration of ZFH ova by FRO was markedly (P 〈 .05) reduced (30.2). Sperm penetration was not significantly correlated with motility or acrosomal integrity before or after incubation, regardless of treatment. These data suggest that the SPA can be used in conjunction with conventional measures of semen analysis in assessing the potential fertilizing capacity of boar sperm and that liquid storage is superior to frozen storage with respect to preserving sperm fertility.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120160302
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  • 15
    Electronic Resource
    Electronic Resource
    Flow cytometry of X and Y chromosome-bearing sperm for DNA using an improved preparation method and staining with Hoechst 33342 (1987)
    New York, NY : Wiley-Blackwell
    Gamete Research 17 (1987), S. 203-212 
    Details
    ISSN: 0148-7280
    Keywords: semen ; sperm sorting ; flow analysis ; cell sorter ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A new and improved method of preparing mammalian spermatozoa for high resolution flow cytometric DNA analysis and flow sorting is described. Ejaculated or cryopreserved sperm were briefly sonicated to remove tails and then stained with Hoechst 33342. This simple procedure was found superior to more severe treatments of dimethylsulfoxide washes, fixation in 80% ethanol, and protease digestion of the sperm membranes and tails by papain. Flow cytometric DNA analyses of sperm samples subjected to varying sonication times indicated that X and Y chromosome-bearing sperm populations could be well resolved with as little as 15-sec sonication. In addition, a comparison of sonicated samples stained with four concentrations of bisbenzimide (Hoechst 33342) or 4′,6-diamidino-2-phenylindole (DAPI) indicated that 2.5 or 5.0 μg/ml of Hoechst was sufficient to resolve the X and Y sperm populations. In order to quantitatively describe the flow cytometric data, several indices (sample quality, orientation and splitting) were developed.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120170303
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  • 16
    Electronic Resource
    Electronic Resource
    Attempted sexing of bovine spermatozoa by fractionation on a Percoll density gradient (1988)
    New York, NY : Wiley-Blackwell
    Gamete Research 20 (1988), S. 83-92 
    Details
    ISSN: 0148-7280
    Keywords: spermatozoal ; separation ; flow cytometry ; semen sexing ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Bovine spermatozoa were fractionated on Percoll density gradients into two major subpopulations of motile spermatozoa and a minor fraction containing mostly nonmotile spermatozoa with abnormal morphology. Fractionation required the addition of bovine serum albumin and a continuous Percoll gradient buffered with sodium bicarbonate. It is postulated that, under suitable ionic conditions, the binding of bovine serum albumin to spermatozoa amplifies subtle differences between subpopulations. These studies were directed toward separating Y- and X-bearing spermatozoa. However, when the subpopulations were evaluated by flow cytometry, their Y:X ratios were similar to that of an unfractionated control.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/mrd.1120200108
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  • 17
    Electronic Resource
    Electronic Resource
    Testicular function of rats following exposure to microwave radiation (1983)
    New York, NY [u.a.] : Wiley-Blackwell
    Bioelectromagnetics 4 (1983), S. 107-114 
    Details
    ISSN: 0197-8462
    Keywords: spermatogenesis ; microwave radiation ; germinal tissue ; Life and Medical Sciences ; Occupational Health and Environmental Toxicology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Physics
    Notes: Male Sprague-Dawley rats were exposed for 6 h per day for nine days to pulse-modulated microwave radiation (1.3 GHz, at 1-μs pulse width, 600 pulses per second). Exposures were carried out in cylindrical waveguide sections at a mean dose rate of 6.3 mW/g; sham controls were treated similarly and received no irradiation. At time periods corresponding to 0.5, 1.0, 2.0, and 4.0 cycles of the seminiferous epithelium, groups of four shamirradiated and four irradiated rats were killed and the testes removed for analysis. Net mass of the testes, epididymides, and seminal vesicles; daily sperm production (DSP) per testis and per gram of testis; sperm morphology; and the number of epididymal sperm were determined. There were no statistically significant differences between the shamirradiated and irradiated groups with respect to any measured variable. In a group of seven surrogate animals of similar body mass, the dose rate of 6.3 mW/g caused a net change in body temperature (via rectal probe) of 1.5 °C.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bem.2250040202
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  • 18
    Electronic Resource
    Electronic Resource
    Acute, whole-body microwave exposure and testicular function of rats (1987)
    New York, NY [u.a.] : Wiley-Blackwell
    Bioelectromagnetics 8 (1987), S. 37-43 
    Details
    ISSN: 0197-8462
    Keywords: neurohormones ; testis ; Life and Medical Sciences ; Occupational Health and Environmental Toxicology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Physics
    Notes: Male Sprague-Dawley rats were exposed for 8 h to continuous-wave microwave radiation (MWR, 1.3 Ghz) at a mean specific absorbed dose rate of 9 mW/g. MWR exposure and sham-irradiation took place in unidirectionally energized cylindrical waveguide sections, within which the animals were essentially unrestrained. The MWR treatment in this setting was determined to yield an elevation of deep rectal temperature to 4.5 °C. The animals were taken for analysis at 6.5, 13, 26, and 52 days following treatment, which corresponded to 0.5, 1, 2, and 4 cycles of the seminiferous epithelium. Net mass of testes, epididymides, and seminal vesicles; daily sperm production (DSP) per testis and per gram of testis; and the number of epididymal sperm were determined. The levels of circulating follicle-stimulating hormone (FSH) and leutinizing hormone (LH) were derived via radioimmunoassay of plasma samples taken at the time of sacrifice. Despite the evident acute thermogenesis of the MWR at 9 mW/g, no substantial decrement in testicular function was found. We conclude that, in the unrestrained rat, whole body irradiation at 9 mW/g, while sufficient to induce evident hyperthermia, is not a sufficient condition for disruption of any of these key measures of testicular function.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bem.2250080106
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