PMID: 22553829May 4, 2012Paper

Tris buffer improves fluorescence yield of ram spermatozoa when evaluating membrane integrity

Microscopy Research and Technique
J L YánizP Santolaria

Abstract

This study was designed to evaluate the effect of various buffers on the fluorescence signal intensity of two fluorochromes (IP and CFDA) when used to assess the membrane integrity of ram sperm. Second ejaculates (18) from nine adult males were collected using an artificial vagina and diluted in either MOPS, TRIS, TES, HEPES, citrate, or phosphate-based extenders. Semen samples were stored at 15°C and the membrane integrity was assessed within the first 24 h of storage. Mean fluorescence intensity (FI) of PI- and CDFA-labeled sperm heads and fluorescence background noise (FBN) were determined quantitatively using Image J software. Fluorescence contrast (FC) was expressed as the difference between FI and FBN. Significantly, higher FI and FC were recorded when TRIS diluent was used, rather than the other diluents, both in the propidium- and fluorescein-labeled cells. The citrate and phosphate-based extenders showed intermediate results of FC between those of TRIS and zwitterionic (MOPS, TES and HEPES) groups for the PI-labeled sperm. However, in the CFDA-labeled sperm, the lower values of FC were obtained in the citrate and phosphate groups due to increased levels of FBN. For the membrane-damaged sperm, fluorescent labeling was l...Continue Reading

References

Jan 1, 1990·Journal of Reproduction and Fertility·R A Harrison, S E Vickers
Feb 1, 1966·Biochemistry·N E GoodR M Singh
Aug 1, 1966·Analytical Biochemistry·D P HanlonE W Westhead
Jul 1, 1982·Journal of Reproduction and Fertility·O A OsinowoL O Eduvie
Aug 1, 1995·Biology of Reproduction·D L Garner, L A Johnson
Jul 1, 1993·Journal of Reproduction and Fertility·J L GattiJ L Dacheux
Jul 31, 2003·Reproduction in Domestic Animals = Zuchthygiene·H Rodríguez-Martínez

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Citations

Jan 9, 2013·Reproduction in Domestic Animals = Zuchthygiene·J L YánizP Santolaria
Mar 8, 2014·Animal Reproduction Science·S Vicente-FielJ L Yániz
Jul 24, 2013·Reproduction in Domestic Animals = Zuchthygiene·J L YánizP Santolaria

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