應用於人工生殖技術之 高泳動力精蟲快速篩選晶片開發

Abstract

體外受精技術(In Vitro Fertilization, IVF)以及單一精蟲微影注射(intracytoplasmic sperm injection)為現今最常使用的人工協助生殖科技，兩項技術都十分強調高品質精子細胞篩選的重要性。 然而，如何將高品質精子從精液當中篩選出來還是一個必須克服的問題。 因此，本研究提出了利用精子本身的運動特性及不同等級的泳動能力來做為高品質精子篩選的方法。精子本身具有泳動能力，佐以實驗證實了具有一定程度泳動能力的精子有朝著流場反方向逆游的特性。利用此一特殊現象以及搭配流道的設計，本生物晶片可直接從未經處理的精夜樣本將高泳動力精子從白血球、脂肪、無泳動力精子以及其他雜質當中篩選出來，並且在短時間之內篩選出大量的高泳動力精子細胞而不需要任何外接電路或是其他類似幫浦等儀器。 本研究利用靜水壓做為穩定的壓力驅動源，整個晶片僅包含了未經處理精液入口以及緩衝溶液入口,而高泳動力精子篩選流道中的流速精準控制在50μm/s，由於每個精子細胞的泳動力皆不同意即抵抗背景流速能力亦不同，而精子的泳動速度約在數微米到數十微米的範圍，如此一來50μm/s高流速流場的配置便可將具有高泳動力的精子分離出來，且佐以實驗證實此機制篩選出的精子活性比例接近百分之百。 此微小、可隨身攜帶、用完即丟的生物晶片可以協助簡化傳統精子篩選繁瑣的步驟並在短時間之內有效的將所需要的高泳動力京子篩選分離出來，以利IVF以及ICSI、或其他相關人工生殖技術的進行，滿足臨床上的需求。

Assisted Reproductive Technology (ART), includes in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), as the two most commonly used technologies to address infertility problems. These approaches reveal the fact that pre-selection of functional sperm is of prime importance to successful fertilization. However, one common obstacle for IVF and ICSI procedures is to select the most competent sperm population from a given semen sample. This thesis enhances motility-based sperm preparation to provide competent sperm for ART. A high throughput microfuidic sperm sorting device that can efficiently separate motile sperms from nonmotile sperms and debris is proposed to fulfill a need in the clinical setting where small amount of qualified sperms need to be sorted. The device isolates motile sperm based on the motility, self-movement, and against-flow phenomenon of motile sperm in a geometrically constrained micro-scaled environment. More specifically, the microfuidic device introduces a flow field for sperms to swim against, and sperms that overcame the flow resistance within a few minutes are propelled along in the microchannels and migrated upstream to the collection region. Hundreds of microchannels are fabricated into the device with a view to increasing the throughput. Hydrostatic pressure was used as the steady driving source by controlling the height of water column to generate the flow field for testing sperm in motility. It has been proven that the hydrostatic pressure and against-flow phenomenon of motile sperm can be used to separating, aligning, and orienting sperm in a microchannel. This microfluidic device is simple to operate, portable and reusable. It can be integrated completely with observation, isolation, and classification of motile sperm in one chip without the need for external pumping system. The microdevice should be applicable to address clinical need of sperm classification and separation for ART.