view - Department of Reproduction, Obstetrics and Herd Health

CHAPTER 3.2
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In veterinary medicine, a number of studies have focussed on counting chambers in
combination with CASA devices. Discrepancies between CASA estimates and the Bürker
haemocytometer have been described for different species (Rijsselaere et al., 2003; Vyt et al., 2004).
To obtain a sharp image of the sperm cells when performing the motility analysis, CASA devices are
primarily used in combination with shallow chambers to keep the moving sperm within focus plane
of the microscope. Therefore CASA systems are frequently used in combination with 20 µm deep
disposable counting chambers. These chambers are most often loaded by capillary force, and such a
capillary flow in a 20 µm chamber follows the laminar Poiseuille flow (Douglas-Hamilton et al.,
2005a). This leads to a transverse lifting force on suspended particles and results in an unevenly
concentration throughout the sample as described by Segre and Silberberg and is, as such, known as
the SS effect (Douglas-Hamilton et al., 2005b). The influence of this SS effect on the distribution
throughout the slide depends on the viscosity of the sample. Correction factors are available and are
based on the time it takes for a chamber to be filled, and as such determined by viscosity of the
loaded sample (Douglas-Hamilton et al., 2005a). Although the SS effect is well documented in
literature, interpretation of the results is not always performed correctly. For example, in a recent
study (Maes et al., 2010), two types of 20 µm Leja slides were compared, where one of the slides
was especially developed to be able to correct for the SS effect. Unfortunately, the results obtained
with the two chambers were compared as such, without applying the correction factor. Therefore it
was faulty concluded that the newly shaped counting chamber was not able to correct for the SS
effect.
CASA instruments were developed to provide a solution for problems linked with subjective
motility analysis. Indeed, subjective assessment of motility is known to be inaccurate, imprecise and
subject to variability (Davis and Katz, 1993; Matson, 1995). However, in order for CASA systems to
contribute to a standardized and objective analysis, uniformity in protocols and settings is required.
In literature, a large variation in technical settings can be found as well as a lack in uniformity for
preparing a sample for motility analysis. It has been demonstrated that the large variety in motility
settings used influences motility outcomes significantly (Hoogewijs et al., 2009). The chambers used
in combination with CASA are very diverse and quite often not clearly described. As such, it is nearly
impossible to compare results between different studies.
The big difference in loading technique and motility outcomes may be attributed by physical
forces on the spermatozoa during loading as postulated by Lenz et al. (2011). When a semen sample
is placed on the loading area of a capillarity filled chamber, the flow that occurs might damage the