Erratum to: 56 SINGLE LAYER CENTRIFUGATION THROUGH PURESPERM® 80 IMPROVES QUALITY OF CRYOPRESERVED DOG SPERMATOZOA

Abstract

Density gradient centrifugation with PureSperm® (PureSperm® 40+PureSperm® 80; Nidacon International, Mölndal, Sweden) has been satisfactorily used to enhance the quality of dog semen samples; however, no studies have been performed on the effect of single layer centrifugation (SLC) with PureSperm® on frozen–thawed dog semen. The aim of this study was to investigate if SLC with PureSperm® 80 can improve the post-thaw semen quality of dog. Semen from 5 dogs was collected by digital manipulation. Two ejaculates from each dog were centrifuged with Tris-based extender, supernatant was removed, and sperm pellet was suspended to a final concentration of 300–400×106spermmL–1 with CaniPROTM Freeze A plus 20% egg yolk at 22°C. Extended semen was cooled to 5°C within an hour and then diluted to a final concentration of 150–200×106spermmL–1 in CaniPROTM Freeze B plus 20% egg yolk at 5°C, loaded in 0.5-mL plastic straws and frozen horizontally in ranks placed 4cm above the surface of liquid nitrogen vapors for 10min, after which they were directly placed in liquid nitrogen. After 24 to 48h of storage, straws were thawed in a water bath at 37°C for 30s. After thawing, semen samples were divided in 2 aliquots: one of them was used as control and the other one was processed by SLC PureSperm® 80. Assessment of sperm motility (assessed by computerized-assisted semen analysis), morphology (Diff-Quick staining), and viability [triple fluorescent stain of propidium iodine/isothiocyanate-labeled peanut (Arachis hypogaea) agglutinin/Rhodamine 123] were evaluated in control and treated semen samples. Data were studied by ANOVA. Results are expressed as mean ± SEM. Significant (P<0.001) differences were found between SLC-treated and control semen for sperm motility (percentage of total motile spermatozoa: 93.65±0.05 v. 83.79±0.13; percentage of progressive motile spermatozoa: 79.38±6.66 v. 54.61±16.11), morphology (86.45±0.01 v. 83.51±0.01), and viability (percentage of viable sperm with an intact acrosome: 58.32±0.04 v. 36.50±0.17; percentage of viable sperm with an acrosome reaction: 2.81±0.01 v. 9.74±0.21). Based on our results, we can conclude that SLC with PureSperm® 80 is an alternative and successful method for improving the quality of frozen–thawed dog spermatozoa, selecting good-quality spermatozoa (motile, morphologically normal, viable, and acrosome intact spermatozoa) from the rest of the semen sample.