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. 2020 Oct 29;10(11):1993.
doi: 10.3390/ani10111993.

Optimization of CASA-Mot Analysis of Donkey Sperm: Optimum Frame Rate and Values of Kinematic Variables for Different Counting Chamber and Fields

Sabrina Gacem  1 , Jaime Catalán  1 , Anthony Valverde  2 , Carles Soler  3 , Jordi Miró  1
Affiliations

Optimization of CASA-Mot Analysis of Donkey Sperm: Optimum Frame Rate and Values of Kinematic Variables for Different Counting Chamber and Fields

Sabrina Gacem et al. Animals (Basel). .

Abstract

In order to optimize the donkey sperm motility analysis by the CASA (Computer Assisted Sperm Analysis)-Mot system, twelve ejaculates were collected from six jackasses. Capillary loaded chamber (CLC), ISAS®D4C depths 10 and 20 µm, ISAS®D4C Leja 20 and drop displacement chamber (DDC), Spermtrack® (Spk) depths 10 and 20 µm were used. Sperm kinematic variables were evaluated using each chamber and a high-resolution camera capable of capturing a maximum of 500 frames/second (fps). The optimum frame rate (OFR) (defined according to curvilinear velocity-VCL) was dependent on chamber type. The highest OFR obtained was 278.46 fps by Spk20. Values for VCL, straight-line velocity (VSL), straightness (STR), amplitude of lateral head displacement (ALH) and beat cross frequency (BCF) were high in DDC and 10 µm depth. In both DDC 10 and 20 µm, the sperm velocities (VCL, VSL, VAP) and ALH values decreased significantly from the centre to the edges, while Wobble and BCF increased. No defined behavior was observed along the CLC. However, all the kinematic variables had a higher value in a highly concentrated sample, in both chamber types. In conclusion, analyzing a minimum of nine fields at 250 fps from the centre to the edges in Spk10 chamber using a dilution of 30 × 106 sperm/mL offers the best choice for donkey computerised sperm motility analysis.

Keywords: capillary loaded chambers; chamber depth; drop displacement chambers; field; frame rate; sperm dilution.

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Conflict of interest statement

The authors declare no conflict of interest. Some of the authors (C.S. and D.B.) are employees for the company that manufacture the CASA-Mot system and the counting chambers used or collaborate through a collaborative agreement between the València University and the company that manufacture some of the components used here. In any case, it does not imply any conflict of interest, because no publicity (direct or indirect) is done in the manuscript. These components were just used in the same way we could use from other brands and the elements studied are not related with the brand but with physic and biological topics.

Figures

Figure 1
Figure 1
The different chambers types and forms used and the filed analyzed. (a): Disposable chamber D4C10, D4C20. (b): Disposable chamber D4C20L and (c): Reusable chambers SpK10, SpK 20. The arrow demonstrates the place of drop deposition. Numbers (1–7) and characters (A, B, C) inside the chambers demonstrate different capture fields.
Figure 2
Figure 2
Sperm curvilinear velocity (VCL. μm/s) obtained at different frame rates in the different counting chambers.
Figure 3
Figure 3
Donkey sperm motility tracks at different frames (25, 50 and 150), exhibiting 4 groups of spermatozoa velocity: rapid (red), medium (green), slow (blue) and static (yellow) in D4C10 chamber, fps = frames per second.
See this image and copyright information in PMC

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