Optimizing Sperm Cell Assessment with the LUNA-FX7™ Automated Cell Counter

Introduction

Sperm cell assessment, also known as a semen analysis, plays an important role in determining male pregnancy potential. While not a direct measure of fertility, male pregnancy significantly influences both human fertility and livestock breeding. Hence, there is high interest in sperm cell assessment.
There are methods available for sperm cell assessment using flow cytometry and image-based analysis tools. Automated cell counters like the LUNA-FX7™ automated cell counter can offer a rapid and convenient methods for assessing sperm samples using images. However, optimized analysis parameters and suitable dyes are required due to the unique morphology and properties of sperm cells compared to mammalian cells.
We conducted a comparative analysis of different fluorescent dyes such as SYBR14, acridine orange (AO), and propidium iodide (PI) to optimize sperm cell analysis using the LUNA-FX7™. This study aims to assess the efficacy of dye combinations and optimize parameters for sperm cell assessment.

Method and Material

Sample Preparation:

The cryopreserved semen samples from Korean cattle were thawed, and the buffer was replaced with PBS. The semen samples were diluted in PBS at a ratio of approximately 1:20 to 1:40, depending on the semen samples.

Dyes:

Fluorescence Staining:

1. Mix:
• 18 μL sperm cells in PBS
• 2 μL of dyes prepared by mixing green and red fluorescent dyes
2. Incubate for 10 min at 37℃
3. Load samples on a desirable slide.
4. Perform analysis using the LUNA-FX7™.