伊红 (Eosin) 又称曙红,属人工合成染料中的呫吨类染料,为桃红色或粉红色的粉末,分子式为C₂₀H₆O₅Br₄Na₂,分子量为691.86;临床上可通过检测精子膜的完整性来评价精子的存活率。
阿拉丁精子活体染色液 (伊红法) 可以常规检测所有精子标本的存活率,通过染料拒染法来鉴别细胞膜完整的精子,从而得出活精子的百分率,该染色液由伊红、去离子水等组成,利用了染料拒染原理,即相关基于损伤的细胞膜,如在非活的(死)细胞上的膜,允许非透过膜性染料可进入膜内染色;而活细胞的细胞膜能够抗拒染料进入,所以产生拒染现象,不着色。该试剂仅适用于科研领域,不适用于临床诊断或其他用途。
操作步骤(仅供参考):
1、取洁净载玻片,滴加新鲜精液和精子活体染色液 (伊红法) 各1滴,混匀。
2、加盖盖玻片,放置30~60s,立即在高倍镜 (可用400×) 下观察。
3、计数200个精子,计算未着色 (活精子) 占200个精子的百分率。
染色结果:
活精子 不着色(白色)
死精子 红色
注意事项:
1、精液标本一旦液化应立即检测精子存活率,最好在30min内,在任何情况下都不能超过1h,以免因脱水或温度变化导致精子失活而使染色检测结果不准。
2、试剂开封后请尽快使用,以防影响后续实验效果。
3、为了您的安全和健康,请穿实验服并戴一次性手套操作。
Eosin, also known as tetrabromofluorescein, is a xanthene dye among synthetic dyes, appearing as a peach red or pink powder with the molecular formula C₂₀H₆O₅Br₄Na₂ and a molecular weight of 691.86. Clinically, sperm viability can be assessed by detecting the integrity of the sperm membrane. Aladdin Sperm Vital Staining Solution (Eosin Method) enables routine viability testing of all sperm samples. It identifies sperm with intact cell membranes via the dye exclusion test, thereby determining the percentage of viable sperm. Composed of eosin, deionized water and other components, this staining solution is based on the principle of dye exclusion: damaged cell membranes, such as those of non-viable (dead) cells, allow impermeant dyes to penetrate and stain the intracellular space, whereas the cell membranes of viable cells can resist the entry of dyes, resulting in the dye exclusion phenomenon with no staining observed. This reagent is for research use only and not intended for clinical diagnosis or any other applications.
Operating Procedures (For Reference Only):
1. Take a clean glass slide, add 1 drop each of fresh semen and Sperm Vital Staining Solution (Eosin Method), and mix well.
2. Cover with a coverslip, let stand for 30–60 seconds, and observe immediately under a high-power microscope (400× recommended).
3. Count 200 sperm cells and calculate the percentage of unstained sperm (viable sperm) among the total 200.
Staining Results
Viable sperm Unstained (white)
Non-viable sperm Red
Precautions:
1. Sperm viability testing should be performed immediately after semen liquefaction, preferably within 30 minutes and no more than 1 hour under any circumstances. Delayed testing may cause sperm inactivation due to dehydration or temperature changes, leading to inaccurate staining results.
2. Use the reagent as soon as possible after opening to avoid compromising subsequent experimental outcomes.
3. For your safety and health, wear a lab coat and disposable gloves during operation.