CAJ | 학술논문

目的对微流控芯片微管道的深度和筛选时间进行优化,构建有效的精子活力筛选器件.方法设置深度分别为25、50、100、200 μm的微管道,在管道入口池分别加入2.5μl小鼠精子悬液,比较不同深度管道的出口池中精子活力和精子相对数量的差异.选取最适深度的芯片用于筛选时间的优化,分别在加入小鼠精子后5、15、30、60 min记录出口池的精子相对数量和精子活力,比较不同筛选时间所得到的精子活力和精子相对数量的差异,以寻找最适的筛选条件.结果深度为25、50、100、200μm的4组管道出口池中精子活力分别为(85.4±2.3)%、(85.8±5.8)%、(87.2±2.8)%、(76.5±2.8)%,差异有统计学意义(F=5.8,P＜0.05),在深度为25～100μm的3组管道之间变化不明显,而在深度为200 μm管道中该指标明显下降;4组不同深度管道出口池中的精子相对数量分别为(5.2±2.0)%、(7.2±2.5)%、(12.3±2.0)%、(7.7±1.1)%,差异有统计学意义(F=6.9,P＜0.05),在25～100 μm范围内随深度的增加而增大,在100μm深度时达到最大值,管道深度增至200 μm时该指标反而下降,综合考虑精子相对数量和活力2项指标,100μm深度的管道为精子活力筛选的最适管道.加入精子后5、15、30、60 min,出口池中精子活力逐渐下降,分别为(99.6±0.7)%、(87.2±2.8)%、(79.3±2.2)%、(62.6±8.0)%,差异有统计学意义(F=37.3,P＜0.01);而出口池中精子相对数量在此过程中提高,分别为(5.8±1.1)%、(10.6±0.9)%、(12.1±1.7)%、(17.9±3.4)%,差异有统计学意义(F=17.8,P＜0.01),加入精子后15～30 min是理想的精子活力筛选时间.结论本研究对微流控芯片管道的深度和筛选时间进行了优化,构建了有效的精子活力筛选器件,为在芯片上实现健康精子的筛选奠定了基础.
목적 대미류공심편미관도적심도화사선시간진행우화,구건유효적정자활력사선기건.방법 설치심도분별위25、50、100、200 μm적미관도,재관도입구지분별가입2.5μl소서정자현액,비교불동심도관도적출구지중정자활력화정자상대수량적차이.선취최괄심도적심편용우사선시간적우화,분별재가입소서정자후5、15、30、60 min기록출구지적정자상대수량화정자활력,비교불동사선시간소득도적정자활력화정자상대수량적차이,이심조최괄적사선조건.결과 심도위25、50、100、200μm적4조관도출구지중정자활력분별위(85.4±2.3)%、(85.8±5.8)%、(87.2±2.8)%、(76.5±2.8)%,차이유통계학의의(F=5.8,P＜0.05),재심도위25～100μm적3조관도지간변화불명현,이재심도위200 μm관도중해지표명현하강;4조불동심도관도출구지중적정자상대수량분별위(5.2±2.0)%、(7.2±2.5)%、(12.3±2.0)%、(7.7±1.1)%,차이유통계학의의(F=6.9,P＜0.05),재25～100 μm범위내수심도적증가이증대,재100μm심도시체도최대치,관도심도증지200 μm시해지표반이하강,종합고필정자상대수량화활력2항지표,100μm심도적관도위정자활력사선적최괄관도.가입정자후5、15、30、60 min,출구지중정자활력축점하강,분별위(99.6±0.7)%、(87.2±2.8)%、(79.3±2.2)%、(62.6±8.0)%,차이유통계학의의(F=37.3,P＜0.01);이출구지중정자상대수량재차과정중제고,분별위(5.8±1.1)%、(10.6±0.9)%、(12.1±1.7)%、(17.9±3.4)%,차이유통계학의의(F=17.8,P＜0.01),가입정자후15～30 min시이상적정자활력사선시간.결론 본연구대미류공심편관도적심도화사선시간진행료우화,구건료유효적정자활력사선기건,위재심편상실현건강정자적사선전정료기출.
Objectiye To optimize the depth of the microchannel and the time point for sperm collection,and improve the efficiency of sperm screening on a microfluidic device. Methods Microchannels with four different depths of 25, 50, 100 and 200 μm were tested. Mice sperm were added to the inlet of the microchannel. The relative quantity and motility of sperm in the outlet were recorded at different collection times, i.e. ,5, 15, 30 and 60 min. Statistical method one-way ANOVA and appropriate post-hoc testing were applied to analyze differences between different groups, and further to select the best-fit depth of the microchannel and the time point for collection. Results In microchannels with depths of 25, 50, 100 and 200 μm, the sperm motilities measured in each outlet were (85.4 ± 2.3)%, (85.8 ± 5.8)%,( 87. 2 ± 2. 8 ) %, (76. 5 ± 2. 8 ) % respectively with statistical significance ( F = 5.8, P ＜ 0. 05 ). No obvious differences were found among 25-100 μm channels, however the motility dramatically decreased in the 200 μm group. The relative sperm quantities were (5.2 ±2.0)%, (7.2 ±2.5)%,(12.3 ±2.0)%,(7. 7 ± 1.1 ) % respectively with statistical significance ( F = 6. 9, P ＜ 0. 05), which increased with channel depth from 25 to 100 μm,while it decreased in the 200 μm channel Taking 2 indexes into account, 100 μmwas the most fit channel depth for sperm motility screening. The sperm motility in the outlet gradually decreased with time. At the time points of 5, 15, 30 and 60 min after adding sperm, the sperm motilities were (99. 6 ±0. 7)%, (87.2 ±2. 8)%, (79. 3 ±2. 2)% and (62. 6 ±8.0)% respectively with statistical significance ( F = 37. 3, P ＜ 0. 01 ). Yet the relative quantities of sperm in the outlet increased almost three times in this process. At the time points mentioned above, the relative quantities of sperm were (5.8±1.1)%, (10.6 ± 0.9)%, (12.1 ± 1.7)%, (17.9 ± 3.4)% respectively with statistical significance ( F = 17.8, P ＜ 0. 01 ). Thus 15-30 min was the ideal screening time. Conclusion An effective microdevice for sperm screening with optimized depth and collection time period is developed,which may contribute significantly for the screening of healthy sperm on microfluidic chips.