view - Department of Reproduction, Obstetrics and Herd Health
view - Department of Reproduction, Obstetrics and Herd Health
view - Department of Reproduction, Obstetrics and Herd Health
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CHAPTER 3.4<br />
3.4.3. Materials <strong>and</strong> Methods<br />
126<br />
3.4.3.1. Stallions <strong>and</strong> semen preparation<br />
The semen used in this experiment was frozen in 0.5 mL straws at two European approved AI<br />
centers in Belgium. The straws were concentrated to contain about 300 × 10 6 spermatozoa per mL.<br />
Four straws per stallion, from the same ejaculate, were thawed in a water bath <strong>of</strong> 38°C for 30s, dried<br />
<strong>and</strong> pooled in a small cup, <strong>and</strong> kept at 38°C. An aliquot <strong>of</strong> 200µL was diluted tenfold with INRA96<br />
(IMV-technologies, L’Aigle, France), a clear semen extender free <strong>of</strong> debris when visualized<br />
microscopically. The samples, diluted <strong>and</strong> undiluted, were incubated at 38°C for 10 minutes prior to<br />
analysis.<br />
3.4.3.2. Analysis with the SQA-Ve<br />
The frozen-thawed semen was analyzed after proper incubation without diluting the thawed<br />
semen according to the manufacturer ‘s guidelines. The semen was aspirated into the specifically<br />
designed capillaries, wiped clean <strong>and</strong> inserted into the device following onscreen instructions. Prior<br />
to analysis, the SQA-Ve preheats the sample once more during 60 seconds after which the analysis<br />
starts automatically. The SQA-Ve does not require parameter settings, the s<strong>of</strong>tware version used was<br />
1.00.43 for experiment 1, <strong>and</strong> 1.00.61 for experiment 2.<br />
When total motility (TM) was equal to or lower than 30%, no actual value is being displayed,<br />
resulting in lack <strong>of</strong> progressive motility (PM) data. However, the number <strong>of</strong> progressive motile<br />
spermatozoa (PMS) per mL remains available. As soon as TM is lower than 10%, PMS is not reported<br />
either. As such, analysis with SQA-Ve can either result in actual values (complete data), in limited<br />
data (TM≤30%, <strong>and</strong> actual value for PMS), or no data at all (TM