Spectrophotometric Analysis of the Resazurin Reduction Test as a Tool for Assessing Canine Semen Quality

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Abstract

The resazurin reduction test (RRT) was subjected to spectrophotometric analysis to evaluate the quality of canine semen. Twenty four samples of canine semen were analysed. The absorption peaks for resazurin and resorufin were determined at 615 and 580 nm, respectively. The RRT ratio (RRTsperm-the ratio for samples containing spermatozoa, RRTplasma-the ratio for samples containing seminal plasma) was calculated by dividing the absorbance at 580 nm by the absorbance at 615 nm. Spearman’s correlation test was used to determine the significance of correlations between the analysed sperm parameters and the results of the resazurin reduction assay. The RRT ratio was highly correlated with sperm motility (r=0.68, P<0.01), progressive sperm motility (r=0.61, P<0.01), the subpopulation of cells with rapid velocity (r=0.72, P<0.01), and the subpopulation of cells with medium velocity (r= -0.54, P<0.05). A negative correlation was observed between the reducing capacity of seminal plasma vs. sperm with plasma membrane integrity (r= -0.60, P<0.01) and sperm with normal morphology (r= -0.58, P<0.01). The RRT test can be used as an additional tool for evaluation of the quality of canine semen.

References
  • 1. Carter R.A., Ericsson S.A., Corn C.D., Weyerts P.R., Dart M.G., Escue S.G., Mesta J.: Assessing the fertility potential of equine semen samples using the reduction dyes methylene green and resazurin. Arch Androl 1998, 40, 59-66.

  • 2. Dart M.G., Mesta J., Cremshaw C., Ericsson S.A.: Modified resazurin test for determining the fertility potential of bovine spermatozoa. Arch Androl 1994, 33, 71-75.

  • 3. Glass R.H., Ericsson S.A., Ericsson R.J., Drovin M.T., Marcoux L.J., Sullivan J.: The resazurin reduction test provides an assessment of sperm activity. Fert Steril 1991, 56, 743-746.

  • 4. Mahmoud A.M., Comhaire F.H., Vermeulen L., Andreou E.: Comparison of the resazurin test, adenosine triphosphate in semen and various sperm parameters. Hum Reprod 1994, 9, 1688-1693.

  • 5. Niżański W., Partyka A., Rijsselaere T.: Use of fluorescent stainings and flow cytometry for canine semen assessment. Reprod Dom Anim 2012, 47, Suppl 6, 215-221.

  • 6. O’Brien J., Wilson I., Orton T., Pognan F.: Investigation of the Alamar Blue (resazurin) fluorescent dye for the assessment of mammalian cell cytotoxicity. Eur J Biochem 2000, 267, 5421-5426.

  • 7. Reddy K.V., Bordekar A.D.: Spectrophotometric analysis of resazurin reduction test and semen quality in men. Indian J Exp Biol 1999, 37, 782-786.

  • 8. Strzeżek R., Filipowicz K., Stańczak M.: Comparison of sperm motility parameters with resazurin reduction assay. Acta Biochim Pol 2012, 59, Suppl 3, 152.

  • 9. Strzeżek R., Koziorowska-Gilun M., Kowalówka M., Strzeżek J.: Characteristics of antioxidant system in dog semen. Pol J Vet Sci 2009, 12, 55-60.

  • 10. Thomas C.A., Garner D.L., DeJarnette J.M, Marshall C.E.: Effect of cryopreservation on bovine sperm organelle function and viability as determined by flow cytometry. Biol Reprod 1998, 58, 786-793.

  • 11. Wang S., Holyoak G.R., Panter K.E., Liu Y., Evans R.C., Bunch T.D.: Resazurin reduction assay for ram sperm metabolic activity measured by spectrophotometry. Proc Soc Exp Biol Med 1998, 217, 197-202.

  • 12. Watson P.F.: Use of Giemsa stain to detect changes in acrosomes of frozen ram spermatozoa. Vet Rec 1975, 97, 12-13.

  • 13. Zalata A.A., Lammertijn N., Christoper A., Comhaire F.H.: The correlates and alleged biochemical background of the resazurin reduction test in semen. Int J Androl 1998, 21, 289-294.

  • 14. Zini A., O'Bryan M.K., Israel L., Schlegel P.N.: Human sperm NADH and NADPH diaphorase cytochemistry: correlation with sperm motility. Urology 1998, 51, 464-468.

  • 15. Zrimsek P., Kurc J., Kosec M., Mrkun J.: Spectrophotometric application of resazurin reduction assay to evaluate boar semen quality. Int J Androl 2004, 27, 57-62.

Bulletin of the Veterinary Institute in Pulawy

The Journal of National Veterinary Research Institute in Pulawy

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IMPACT FACTOR 2016: 0.462

SCImago Journal Rank (SJR) 2016: 0.235
Source Normalized Impact per Paper (SNIP) 2016: 0.364

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