Semen Extension and Cryopreservation Flashcards
Dilute semen in ______ to maintain sperm viability
Buffered extenders
_______ varies per species
Semen extension
Extenders differ in composition depending upon the _____,_____,______and ______
species,
intended use,
storage temperature,
the desired duration of storage
Buffers in extenders modulate the change in extender pH due to _____________
metabolic products of stored spermatozoa
Extenders contain freezing point depressing compounds, which ________,_________, and _________
inhibit ice formation,
dehydrate cells,
and protect cellular structures
Extenders maintain sperm viability at ________and control bacterial growth
20 deg. Celsius
Bovine: (Procedure 1)
To pass for processing, semen samples should have an initial minimum ________progressive
motility and ______ normal morphology
minimum 75% progressive
motility and,
70% or greater normal morphology
Bovine: (Procedure 2)
Semen aliquot is diluted 100-fold in __________, and ______are added for pathogen control
saline solution and atbcs are added for pathogen control
Bovine: (Procedure 3)
__________ is incubated prior to extension since the activity of _____is hindered in the presence of milk or egg yolk in semen extenders
Atbc-semen mixture is incubated prior to extension since the activity of atbcs is hindered in the presence of milk or egg yolk in semen extenders
Bovine: (Procedure 4)
Semen is then added to _________, then placed in an __________
placed into a 5 deg. celsius cold cabinet, cooled for several hours
Semen is then added to isothermal extender, then placed in an isothermal water jacket
placed into a 5 deg. Celsius cold cabinet, cooled for several hours
Bovine: (Procedure 6)
Semen is then diluted to final desired concentration with ________, packaged
in _______ plastic french straws, sealed, and frozen for 10 minutes in _________ at a starting temperature of ________
Semen is then diluted to final desired concentration with glycerolated extender, packaged in 0.5 mL plastic french straws, sealed, and frozen for 10 minutes in static nitrogen vapor at a starting temperature of -160 deg. celsius
Bovine: (Procedure 5)
After initial cooling, samples are ___________________
removed from water jackets and maintained at 5 deg. celsius
Bovine: (Procedure 7)
After freezing, the straws are immersed into _______ until the time of _______, through placing it into a ________water bath for 30-60 seconds
After freezing, the straws are immersed into liquid nitrogen until the time of thawing, through placing it into a 35 deg. water bath for 30-60 seconds
Bovine: (Procedure 9)
Post-thaw analysis minimum standards:
1. Motility
2. Spermatozoal progressive linear motility
3. Intact acrosomes
Post-thaw analysis minimum standards:
1. Motility: 25% (0hrs)- 15% (2hrs)
2. Spermatozoal progressive linear motility: 3-2
3. Intact acrosomes: 60-40%
Bovine: (Procedure 8)
All _________are performed at 35 deg. celsius
post-thaw evaluations
Equine (Procedure 1):
Stallion semen may be diluted with _____ or _____, then used immediately or cooled to ____and stored for AI of mares within _________
Stallion semen may be diluted with egg-yolk buffer or milk solids-glucose extenders, then used immediately or cooled to 5°C and stored for AI of mares within 24 to 48 hours of ovulation
Equine (Procedure 2):
Semen may be centrifuged to_________
remove seminal plasma to concentrate sample
Equine (Procedure 3):
Soft pellet is then extended at 20°C in a_____________ medium containing ______
sodium-citrate-EDTA lactose-egg yolk medium containing cryoprotectant, glycerol
Equine (Procedure 4):
Final extended semen should have a concentration sufficient to provide ________, normal cells after thawing
at least 200 x 106 motile
Equine (Procedure 5):
Semen is then packaged into plastic straws ranging from __________
0.5 mL to 5.0 mL
Equine (Procedure 6):
Freezing is accomplished in a ______ or ______ nitrogen vapor unit, then immediately immersed in ___________
Freezing is accomplished in a passive or mechanically forced nitrogen vapor unit, then immediately immersed in liquid nitrogen for storage at -196°C
Ovine (Procedure 1):
Ram semen can be extended for fresh insemination, _____________ or ________________
cooled for short term storage (24-48hours) or stored frozen for long-term preservation
Ovine (Procedure 2):
Extender
tris-glucose (1 part semen / 2parts extender)
Ovine (Procedure 3):
Thawing
place straws in 35°C water for at least 30 seconds
Caprine (Procedure 1):
Goats inseminated with ___________
fresh extended, cooled, or frozen semen
Caprine (Procedure 2):
Extender
milk or tris-citrate-egg yolk (4-5%) extenders, glycerol
Caprine (Procedure 3):
Initial colling is to _____ in _____
Initial colling is to 5°C in non-glycerolated extender
Caprine (Procedure 4):
_________ is then added slowly and the mixture us allowed to equilibrate over _____ before freezing in nitrogen vapor and storage at ______
Glycerolated portion is then added slowly and the mixture us allowed to equilibrate over 2 hours before freezing in nitrogen vapor and storage at -196°C
Caprine (Procedure 5):
Thawing
35°C, 30 seconds before insemination
Porcine (Procedure 1):
Collected ejaculate is added to a ______and cooled to _______
Collected ejaculate is added to a predilution extender and cooled to 18°C over 4 hours
Porcine (Procedure 2):
Semen centrifuged for ______ at _____and the pellet resuspended in ________________
Semen centrifuged for 10 minutes at 18°C and the pellet resuspended in a cooling diluter containing egg yolk
Porcine (Procedure 3):
Semen cooled from _________
18°C to 5°C
Porcine (Procedure 4):
Before freezing, semen is diluted to a final concentration of __________ in a ________, then frozen in 5 mL plastic tubes
1.2 x 109 spermatozoa/mL in a deep-freeze diluter at 4% glycerol, then frozen in 5 mL plastic tubes
Porcine (Procedure 5):
Thawing
52°C for 50 seconds
