title: Comparison of Cryopreservation Protocols (Single and Two-steps) and Thawing (Fast and Slow) for Canine Sperm.
abstract: In addition to the existence of several cryopreservation protocols, no systematic research has been carried out in order to confirm the suitable protocol for canine sperm. This study aims to assess the effect of adding 5% glycerol during cryopreservation at 37°C (one-step) and 5°C (two-steps), in addition of testing two thawing protocols (37°C for 30 seconds, and 70°C for 8 seconds). We used 12 sperm samples divided into four experimental groups: Single-Step - Slow Thawing Group; Two-Step - Slow Thawing Group; Single-Step - Fast Thawing Group; and Two-Step - Fast Thawing Group. Frozen-thawed samples were submitted to automated analysis of sperm motility, evaluation of plasmatic membrane integrity, acrosomal integrity, mitochondrial activity, sperm morphology, sperm susceptibility to oxidative stress, and sperm binding assay to perivitellinic membrane of chicken egg yolk. Considering the comparison between freezing protocols, no statistical differences were verified for any of the response variables. When comparison between thawing protocols was performed, slow thawing protocol presented higher sperm count bound to perivitelline membrane of chicken egg yolk, compared to fast thawing protocol. Regardless of the freezing process, the slow thawing protocol can be recommended for the large scale cryopreservation of canine semen, since it shows a consistent better functional result.
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Maíra M
authors.initials: MM
authors.lastname: Brito
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Cristina F
authors.initials: CF
authors.lastname: Lúcio
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Daniel S R
authors.initials: DS
authors.lastname: Angrimani
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: João Diego A
authors.initials: JD
authors.lastname: Losano
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Andressa
authors.initials: A
authors.lastname: Dalmazzo
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Marcílio
authors.initials: M
authors.lastname: Nichi
authors.affiliation: a Department of Animal Reproduction, School of Veterinary Medicine and Animal Science , University of São Paulo , São Paulo , Brazil.
authors.forename: Camila I
authors.initials: CI
authors.lastname: Vannucchi
mesh_terms: D000818:Animals; D002645:Chickens; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D003451:Cryoprotective Agents / Q000494:pharmacology*; D004285:Dogs / Q000502:physiology*; D004530:Egg Yolk; D005990:Glycerol; D008297:Male; D012661:Semen / Q000502:physiology; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology; D013997:Time Factors
chemical_list: D003451:Cryoprotective Agents; D005990:Glycerol
keywords: Dog; sperm cryopreservation; thawing