Cryopreservation of cod semen

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Igor Babiak, Oddvar Ottesen and Vegard
Marschhauser
Bodø University College Department of Fisheries
and Natural Sciences Bodø, Norway
Cryopreservation of cod semen
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1. Pre-selection of extenders

• Good extender
• Does not activate spermatozoa movement
• Protection of cells against freezing/thawing
  damage
• Not too toxic to suspended cells

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Methods
Sperm loading French straws 0.5 ml Freezing LN2
vapours Storage LN2 (-196 C) Thawing water bath
(40 C) and then release to Hanks Balanced Salt
Solution Motility examination immediately after
thawing and 30 min after thawing
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Methods
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Spermatozoa motility
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Spermatozoa motility
MOT motile spermatozoa () VCL curvlinear
velocity (µm/s), actual velocity along the
trajectory VSL straight line velocity (µm/s),
straight line distance between start and end
points, divided by the time of the track LIN
linearity, VSL/VCL ()
Actual trajectory of motile spermatozoa
Spermatozoa moving out of the area (excluded from
the analysis)
Static spermatozoa (inmotile)
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Results
20 extenders (5 diluents x 4 cryoprotectants)
examined
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Results
The best post-thaw motility (60 of motile
spermatozoa) Extender composed of HBSS hens
egg yolk (10) glycerol (10)
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2. Selecting extenders
Correlations Post-thaw motility Fertilization
0.83 Post-thaw motility survival at day 10
0.82 Fertilization Survival at day 10 0.99
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3. Effect of individual male
Correlations Post-thaw motility Fertilization
0.75 Post-thaw motility survival at day 10
0.73 Fertilization Survival at day 10 0.99
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4. Individual male, type of cryoprotectant and
sperm-to-egg ratio
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4. Individual male, type of cryoprotectant and
sperm-to-egg ratio
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5. Type of cryoprotectant and sperm-to-egg ratio
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5. Type of cryoprotectant and sperm-to-egg ratio
1 straw 0.5 mL for c. 1,000 eggs