Htr ivos 12

Sperm motility and motion parameters were assessed with the CASA system (HTR-IVOS 12.3, Hamilton Thorne Biosciences, MA, USA) using the software settings described in a previous study [21 (link)]. The sperm parameters assessed by the CASA system included TMOT (%), PMOT (%), VSL (μm/s), VAP (μm/s), VCL (μm/s), STR (ratio of VSL/VAP × 100%), LIN (ratio of VSL/VCL × 100%), amplitude of lateral head displacement (ALH, μm) and beat cross frequency (BCF, Hz).

The Computer-Assisted Sperm Analysis (CASA) system (HTR-IVOS 12.3, Hamilton Thorne Biosciences, Beverly, MA, USA) was used to monitor sperm motility and velocity parameters [14 (link)]. The software settings used for the CASA system were those recommended by the manufacturer for analysis of boar spermatozoa. The CASA system measured the total motility (TMOT, %), progressive motility (PMOT, %), velocity straight line (VSL, µm/sec) and the velocity average path (VAP, µm/sec).

Sperm motility and motility parameters were evaluated using the computer-assisted sperm analysis (CASA) system (HTR-IVOS 12.3; Hamilton Thorne Biosciences, Beverley, MA, USA). Before the analysis, sperm samples were diluted 1:4 with phosphate-buffered saline (PBS) to obtain 20–30 × 10⁶ spermatozoa/mL, and heated at a temperature of 37 °C for around 10 min (Thermo Block TDR-120, Göttingen, Germany). Then, aliquots of sperm samples (5 μL) were placed in a pre-warmed Makler counting chamber (Sefi-Medical Instruments Ltd., Haifa, Israel) and evaluated at 37 °C. The following parameters were analyzed: total motility (TMOT, %), progressive motility (PMOT, %), velocity average pathway (VAP, μm/s), straight line velocity (VSL, μm/s), curvilinear line velocity (VCL, μm/s), amplitude of lateral head displacement (ALH, μm), beat cross frequency (BCF, Hz), linearity coefficient (LIN, %), and straightness (STR, %).
In the CASA analysis, sperm parameters were measured with the following settings (according to the Hamilton Thorne technical guide v. 12.3 for gazelle/deer): frame acquired—60, frame rate—60 Hz, minimum cell contrast—60, minimum cell size—5 pixels, straightness threshold—80%, low VAP cut-off—21.9 μm/s, low VSL cut-off—6.0 μm/s. A minimum of five fields per sample were assessed, with approximately 200 spermatozoa per field.