New techniques for the assessment of canine semen quality: A review T. Rijsselaere * , A. Van Soom, S. Tanghe, M. Coryn, D. Maes, A. de Kruif Department of Reproduction, Obstetrics and Herd Health, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium Abstract Until recently, canine semen assessment was routinely performed by conventional light micro- scopic techniques. The limitations of these methods include subjectivity, variability, the small number of spermatozoa analyzed, and poor correlation with fertilizing potential. The last decade, several new in vitro techniques have been introduced for canine semen assessment that enable a more detailed evaluation of several sperm characteristics. Numerous fluorescent staining techniques have been developed for the evaluation of specific sperm characteristics and functions, including plasma membrane integrity, capacitation status and the acrosome reaction. By combining fluorescent stains, several functional sperm characteristics can be assessed simultaneously. Moreover, by means of flow cytometry, large numbers of fluorescently labelled spermatozoa can be analysed in a short interval. Following thorough standardization and validation, computer-assisted sperm analysis systems provide objective and detailed information on various motility characteristics and morphometric dimensions that cannot be identified by conventional light microscopic semen analysis. In vitro assays, evaluating the capacity of canine spermatozoa to bind to the zona pellucida or oviductal explants, or to penetrate the oocyte, provide additional information on canine gamete interaction that may be useful in predicting the fertilizing potential of spermatozoa. Although substantial improve- ments have been made in canine semen assessment, surprisingly few parameters were correlated with in vivo fertility. Therefore, further research is required to determine which sperm characteristics are of clinical value for predicting the in vivo fertility in dogs. # 2005 Elsevier Inc. All rights reserved. Keywords: Dog semen; Sperm quality; Fluorescent staining; CASA; Zona pellucida binding www.journals.elsevierhealth.com/periodicals/the Theriogenology 64 (2005) 706–719 * Corresponding author. Tel.: +32 9 264 75 64; fax: +32 9 264 77 97. E-mail address: tom.rijsselaere@UGent.be (T. Rijsselaere). 0093-691X/$ – see front matter # 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.theriogenology.2005.05.021