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Development of a protocol for multiple staining with fluorochromes to assess the functional status of boar spermatozoa
Author(s) -
Bussalleu Eva,
Pinart Elisabet,
Yeste Marc,
Briz Mailo,
Sancho Silvia,
GarciaGil Nuria,
Badia Elena,
Bassols Judit,
Pruneda Anna,
Casas Isabel,
Bonet Sergi
Publication year - 2005
Publication title -
microscopy research and technique
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 118
eISSN - 1097-0029
pISSN - 1059-910X
DOI - 10.1002/jemt.20246
Subject(s) - boar , sperm , acrosome , andrology , staining , biology , semen , propidium iodide , anatomy , biochemistry , genetics , medicine , apoptosis , programmed cell death
The aim of this study was to design a simple and reliable method for the simultaneous evaluation of the nucleus, the acrosome, and the mitochondrial sheath of boar spermatozoa. Sperm samples coming from healthy and sexually mature Pietrain boars were incubated with two nuclear fluorochromes—bis‐benzamide specific for viable cells, and propidium iodide specific for nonviable cells—the fluorochrome Mitotracker® Green FM specific for functional mitochondria, and the lectin Trypsin inhibitor from Soybean (SBTI) conjugated with the fluorochrome Alexa Fluor® 488 specific for proacrosin. The results obtained from assessing the functional status of the spermatozoa using fluorochromes were compared with the conventional sperm parameters of sperm vitality using the eosin exclusion test (EE test), and sperm motility and morphology using the computer‐assisted semen analyzer SCA®2002Producció. Applying the multiple staining test, it was found that the frequency of viable spermatozoa with intact acrosome and intact mitochondria was not different from the frequency of viable spermatozoa obtained with the EE test, and also correlated positively with the frequency of motile spermatozoa and the frequency of mature spermatozoa. Therefore, this technique is useful to characterize the status of boar spermatozoa by assessing the nuclear, acrosomal, and mitochondrial integrity. Moreover, it provides reliable diagnostic information about the fertility potential of boars. Microsc. Res. Tech. 68:277–283, 2005. © 2005 Wiley‐Liss, Inc.