The Use of Different Staining Methods in the Evaluation of Frozen-Thawed Cauda Epididymal Ram Sperm Morphology

Abstract

Sperm morphology evaluation is an important parameter for determining the quality of semen and predicting fertility in rams. Different staining methods have been developed to detect the morphological status of sperm, but there is no optimized protocol, especially for animals yet. This study was designed to compare the results using SpermBlue®, Diff-Quick, and Coomassie Blue stain in the morphological evaluation of epididymal ram semen. In the study, samples collected from a Bafra ram (epididymal sperm/ram) known to have a good breeding history were diluted with Tris-based diluent and frozen. After thawing for each straw, three semen smears were made and stained with SpermBlue®, Diff-Quick, and Coomassie Blue. All morphological parameters were evaluated using a light microscope. 100 spermatozoa were examined randomly and classified according to their characteristics for each slide. In identifying morphological abnormalities, the staining protocols have compared amongst themselves, and no significant difference between Diff-Quick and SpermBlue® staining methods was observed. However, significant differences were observed in midpiece abnormalities when SpermBlue® and Coomassie-Blue staining methods were compared, while significant difference was found in total abnormality in SpermBlue® and Diff-Quick staining comparison (P <0.05). As a result, all staining methods evaluated can be easily optimized for laboratory conditions and used in the morphological analysis of ram semen. © 2022 Asociacion Peruana de Reprouduccion Animal. All rights reserved.