201226046 六、發明說明： 【發明所屬之技術領域】 本發明係關於一種血液分離方法，特別係關於一種利 用掃流原理之血液分離方法。 【先前技術】 血液擔負生物體内氧氣及能量傳輸的重要工作，係為 生命系統正常運作不可或缺的元素之一。基於此點，臨床 治療及醫療檢驗領域均發展出各式以血液為中心的應用 技術，希望透過正確且迅速地取得/分析血液中所含的細胞 或物質，進一步解決病痛帶來的不適，提昇人類生活品 質。在此理念下，作為眾多進階應用基礎的血液分離或過 濾技術一直被視為重點發展的項目之一。 目前，已知分離血細胞與a衆的技術主要可分成四大 類。第一種為離心技術，透過灰細胞比重較大的特性，將 全血血液置於試管中，以高速旋轉產生的離心力，達成將 血細胞與血漿分離的目標。然而，由於部分血細胞之密度 及沈澱速度與血漿成分非常接近，甚至相互重疊，導致離 心技術在某些特殊應用上無法提供所需的純度。再者，此 種技術通常必須要消耗大量的樣本及藥品，且處理上耗時 甚長，整體成本偏高。 其二係以介電泳力為基礎原理，搭配微機電製程，以 特殊的儀器設備提供非均勻之交流電場，藉由血細胞與血 漿間不同的導電度和介電常數，使兩者產生分離效果。但 201226046 此技術目前仍受到步驟繁複以及所需設備造價較昂貴等 條件限制，應用範圍略微狭隘。 非接觸式高波長雷射係近年來發展出的新技術，其係 利用相反方向光壓形成一穩定的能量阱，以钳住微小粒 子。雖然，目前學說理論認為將此種工具方法用於分離血 液將具備非接觸性、非侵入性等極佳的優點，讓非接觸式 高波長雷射在此領域中受到非常大的期待，但是實際產業 運用上還是難以克服設備成本過高的問題，推展遭遇相當 •困難。 深層過濾係四種技術中應用為較為普遍者，透過驅動 血液以垂直方向流通過多層薄膜結構，使其中的血細胞等 -固體物質戴留在薄膜表面。然而，此種技術最大的問題在 於薄膜表面的遽餅形成’尤其隨通入血液量的增加，遽餅 厚度會無可避免的對應增加’使得每作用一段時間就必須 進行沖洗移除，耗費相當多的人力。除此之外，更重要的 馨 是’深層過濾、往往會對血液中的血細胞產生較大的壓力， 使得過濾過程中發生溶血現象，致使分離作業失敗。 综合上述，明顯可見習知分離血細胞取得血漿的技術 難免有樣本消費量大、儀器設備昂貴不符合應用成本、作 業耗時長及/或製程步驟繁瑣等問題，因此，如何提供一種 血液過遽方法’其不需猎助面成本的设傷儀器，因此呈有 較佳的應用彈性’並且反應迅速，消耗樣品量少，操作上 較符合經濟效益’更重要的是能有效避免濾餅形成及溶血 現象產生’已成為一項重要的課題。 201226046 【發明内容】 有鑑於上述課題，本發明之目的為提供一種血液分離 方法及避免阻塞及溶血之血液分離方法，其不需藉助高成 本的設備儀器，因此具有較佳的應用彈性，並且反應迅 速，消耗樣品量少，操作上較符合經濟效益，更重要的是 能有效避免濾餅形成及溶血現象產生。 為達上述目的，依據本發明之一種灰液分離方法包括 以下步驟：提供一具有複數孔洞之薄膜；驅動一血液流 動，且於接觸薄膜時，實質上與薄膜相對平行移動；以及 收集血液通過薄膜之孔洞之部分。其中，血液通過薄膜之 礼洞之部分係為ϋ。....... 在本發明一實施例中，本發明血液分離方法更包括藉 由薄膜阻擋血液内之血細胞，且血液之其他部分通過薄膜 之孔洞之一步驟。 在本發明一實施例中，血液係連續地提供，而透過驅 動連續接觸薄膜。 在本發明實施例中，薄膜具有一第一表面及一第二表 面。第一表面具有複數弧面，孔洞設置於該等弧面之間， 且第二表面對應孔洞，具有複數凹陷區。 在本發明實施例中，孔洞之孔徑範圍係為1微米至50 微米。在本發明一實施例中，孔洞之孔徑範圍係為1微米。 在本發明實施例中，薄膜之材料包括金屬及/或合金。 在本發明一實施例中，薄膜之材料為合金。 在本發明實施例中，血液係由一驅動單元驅動。其 201226046 中，驅動單元係為幫浦、抽吸器或其組合。在本發明一實 施例中，血液係由一微量幫浦驅動。 為達上述目的，依據本發明之一種避免阻塞及溶血之 血液分離方法包括以下步驟：提供一具有複數孔洞之薄 膜；驅動一血液流動，且於接觸薄膜時，實質上與薄膜相 對平行移動；以及收集血液通過薄膜之孔洞之部分。其 中，血液通過薄膜之孔洞之部分係為血漿。 在本發明一實施例中，本發明避免阻塞及溶血之血液 分離方法更包括藉由薄膜阻擋血液内之血細胞，且血液之 其他部分通過薄膜之孔洞之一步驟。 在本發明實施例中，薄膜具有一第一表面及一第二表 面。第一表面具有複數弧面，孔洞設置於該等弧面之間， 且第二表面對應孔洞，具有複數凹陷區。 承上所述，因依據本發明之血液分離方法與避免阻塞 及溶血之血液分離方法係使血液以實質上平行的方式流 過具有孔洞的薄膜，再透過孔洞的尺寸限制，將血液中的 血細胞截留於薄膜之一側，從而與例如血漿等液態成分分 離。其中，重要的是，依據掃流原理，由於血液係與薄膜 相對平行移動，故利於形成平行剪切應力，如此，能有效 降低血液分離時濾餅形成的問題，避免孔洞阻塞以及溶血 現象，影響血液分離作業的成功率及效率。 與習知技術相較，本發明之血液分離方法與避免阻塞 及溶血之血液分離方法不需藉助成本昂貴的儀器設備，而 簡單結合例如可供液體流動的流道以及具有適當孔徑孔 7 201226046 洞的薄膜即可運作，顯著地降低使用門檻，適合推廣至各 等級的醫療院所或研究單位使用。再者，本發明方法具有 適於應用在生物晶片的特性，不僅便於攜帶、運送，且能 以少量的樣本完成分析與檢測，操作上較符合經濟效益。 更重要的是，習知技術中常見的濾餅形成及溶血現象等副 作用均可透過血液在流動時形成的剪切應力加以消除，避 免時常需要人工清理的問題。 [實施方式】 以下將參照相關圖式，說明依本發明較佳實施例之血 液分離方’法與避免阻塞及溶血之血液分離方法，其中相同 的元件將以相同的元件符號加以說明。 圖1為依據本發明第一實施例之血液分離方法的步驟 流程圖。請參考圖1所示，在本實施例中，血液分離方法 包括以下步驟：提供一具有複數孔洞之薄膜（S11);驅動 一血液流動，且於接觸薄膜時，實質上與薄膜相對平行移 動（S13);以及收集血液通過薄膜之孔洞之部分（S15)。 其中.，血液通過薄膜之孔洞之部分係為血漿。在此，本實 施例之血液分離方法係用以將血液中的血漿與血細胞分 離，其中，血細胞係例如為紅血球、各種白血球及/或血小 板之統稱。 為使本實施例各步驟的相關細節更為清楚明瞭，以下 將配合一裝置為例，先清楚介紹該裝置之結構與組成，進 而以此為基礎，說明如何於該裝置上實施本發明方法。然 201226046 而，特別需要提出的是，以下所舉實施例中的裝置僅係為 方便說明使用，並非用以限制本發明。當然，其他能與本 發明方法配合之裝置或系統亦可應用於實現本發明技術 内容，並不限於任何特定之結構。 圖2為應用本發明第一實施例之血液分離方法之裝置 的示意圖，而圖3為圖2所示之裝置的分解示意圖。請同 時參考圖2及圖3所示，在本實施例中，應用血液分離方 法的裝置2可以為一微型裝置，例如微型生物晶片，其外 • 觀呈一薄型長方體。就細部結構而言，裝置2可包括一包 覆殼體21、一流道22、一收集槽23以及一過濾用的薄膜 24。其中，包覆殼體21可更'―進一步具有一上流道部211 / 及一下收集部212。上流道部211及下收集部212可以為 一體成形或分離的兩塊體，且兩者尺寸大小可以相同或不 同。在本實施例中，上流道部211與下收集部212係為獨 立製成的兩部分，但長度、寬度及高度均分別為4公分、 0 2.5公分及0.5公分。上流道部211與下收集部212在形成 後，透過密封結合形成包覆殼體21。當然，除上述實施例 外，兩者的尺寸亦可隨應用所需進行調整，非為限制條件。 包覆殼體21及其上流道部211與下收集部212的材 質可例如為聚曱基丙烯酸曱酯（Polymethylmethacrylate, PMMA )、聚雙甲基石夕氧烧(Poly-dimethylsiloxane, PDMS )、環氧樹脂（Epoxy )、金屬或玻璃，當然，其他具 備較佳機械強度以及高生物相容性特性的高分子材料亦 可應用之。在本實施例中，包覆殼體21及其上流道部211 9 201226046 與下收集部212的材質係以聚雙甲基矽氧烷（PDMS)為 例，其不具細胞毒性，適合作為與活體生物樣本接觸的材 料。另外，聚雙甲基矽氧烷還具有透光性佳，方便觀測等 特性。 流道22、收集槽23以及薄膜24均容置於包覆殼體 21。其中，流道22設置於上流道部211，收集槽23設置 於下收集部212,且薄輯24夾設於流道22及收集槽23之 間。流道22係為連續的液體通路，且除底側係為開放， 其餘上側及左右兩側共三面則係為封閉。流道22兩端對 外開口，例如本實施例中外露在包覆殼體21之一表面213 而呈圓形的開口。流道22的開口即分別為液體輸入的流 入口 221及輸出的流出口 222，其可分別連接例如鐵氟龍 (polytetrafluoroethene, PTFE)橋接頭，以便再與注身十針 筒連結，供血液輸入時使用，同時避免外漏的發生。流道 22的整體路徑並不限定形狀，在此係以螺旋狀或渦狀為 例，其可在一定的面積範圍内，形成距離較長的液體通 路，有利於血液分離作業發揮較佳的功效。然而，在其他 實施例中，流道22亦可以呈簡單的直線或鋸齒狀，端視 血液分離需求以及血凌中各成分的比例決定，而能確實將 血液中固態與液態物質分離為原則。 在本實施例中，流道22係以灌膠成形的方式設置於 包覆殼體21之上流道部211。詳細而言，可選用具有流動 性膠體性質的聚雙曱基矽氧烷覆蓋於母模，待膠體凝固 後，將聚雙曱基矽氧烷形成的部分與母模分離，即可形成 201226046 設置有流道22的包覆殼體21之上流道部211。圖4為圖 3所示之流道的放大示意圖。請參考圖4所示，流道22係 設置於上流道部211之中心位置，且流道22整體（以下 以掃流區稱之）概略呈螺旋狀或渦狀。掃流區直徑dl約 0.8公分。流道22的通道寬度d2約為50微米，流道的結 構高度約為100微米 '當然，上述尺寸均非本發明之限制 性條件，而可隨實際應用需要調整，在其他實施例中，流 道22的通道寬度d2可以由約50至500微米，例如為100 或200微米，而流道22的結構高度可以由約10至500微. 米，例如20或200微米。另外，在流道22形成的過程中， 同時透過例如模具的設計使包覆殼體一21之上流道部211 形成對外開口的流入口 221與流出口 222，在此，兩者均 為細長管體（圖4中未示）。 請參考圖3所示，本實施例之收集槽23亦可以依據 上述相同方式而設置於下收集部212,且對應流道22設置 於薄膜24之另一側。其中，收集槽23可以為一直徑0.8 公分之圓形凹槽，且結構高度為200微米。當然，收集槽 23的尺寸亦可隨例如輸入血液的量而調整，在其他實施例 中，其結構高度可介於10至500微米，例如，結構高度 可以為20或200微米。 在本實施例中，掃流區、收集槽23以及薄膜24三者 的中心可例如實質上重疊，且薄膜24夾設於流道22以及 收集槽23間，換言之，薄膜24亦係夾設於上流道部211 及下收集部212間。薄膜24的尺寸及構型並無特別限制，s 11 201226046 原則上係以配合掃流區為原則。 圖5a為圖2所示之薄膜的部分放大示意圖，而圖5b 為圖5a所示之薄膜於剖面線B-B位置的示意圖。請同時 參考圖5a及圖5b所示，在本實施例中，薄膜24可為直徑 1公分的圓形薄膜，其上具有複數孔洞241。薄膜24具有 一第一表面242及一第二表面243，其分別為薄膜24之上 表面及下表面。在本實施例中，巨觀下觀察之薄膜24的 第一表面242及第二表面243均近似平整表面，而無凹凸 起伏變化（如圖3所示.）。然而，請參考圖5b所示，若進 一步以微觀角度觀察，則第一表面242具有複數向上凸起 的弧面244，而孔洞241設置於弧面244之間，>且第二表 面243對應孔洞241，具有複數凹陷區245。 詳細再說明上述結構。請同時參考圖5a及圖5b所示， 在本實施例中，弧面244可例如為圓弧面，且包括頂端及 底端，底端設置於孔洞241的周邊。凹陷區245可以為圓 形，且面積大於孔洞241之面積。此外，孔洞241係為連 通薄膜24之第一表面242至第二表面243的圓形穿孔， 其直徑大小可例如為1微米。然在本發明其他實施例中， 孔洞241亦可以為其他形狀及/或尺寸，例如由1至50微 米，原則上係依據血液分離所欲達成的效果而定。舉例而 言，以完全分離血液中的血細胞為例，孔洞241的較佳直 徑為1微米。 薄膜24可以為金屬及/或合金材質，其製造可以透過 包括微影黃光製程與電鑄（electroforming)等數個步驟。 12 201226046 首先，可先提供一基板，再於基板之一表面上形成一光阻 層。接續，於光阻層形成複數個光阻圖樣·，並顯露於基板 之部分表面。設置一光罩於光阻層上方，其中，光罩具有 複數個相對應光阻圖樣之圖樣。其後，進行曝光及顯影， 以形成本實施例所不之圓形光阻圖樣。利用電禱法將一合 金層形成於光阻圖樣及基板上。最後，將合金層與光阻圖 樣及基板分離開來，即形成可供本實施例方法應用之分離 血液之薄膜24。 • 圖6為圖2所示之裝置於剖面線A-A位置的示意圖。 請參考圖6所示，在本實施例中，由於流道22係實質上 平行疊設於薄膜24之第一表面242上，因此當血液BL通 入並流過流道22時，必然會有部分期間與薄膜24接觸， 且係以實質上平行薄膜24的方式流過薄膜24上方。流道 22藉由本身底側為開放的構型，而與收集槽23透過薄膜 24之孔洞241相連通，是以，當血液BL於流道22中流 0 動時，會受到重力作用而由第一表面241的弧面244流至 孔洞241 (圖中因為巨觀角度，故仍呈平整狀，詳細結構 可參照圖5b)，使血液BL中例如血漿之液態物質的部份 順利通過孔洞241，至於其中血液BL中所含的例如微粒 或顆粒大小超過孔洞241尺寸的固體物質均會被孔洞241 截留，達到血液分離過濾的效果。 值得特別說明的是，應用本發明之血液分離方法之裝 置2的結構，可使血液BL符合掃流過濾的原理，產生平 行剪切應力，從而掃除過濾過程中血細胞堆積於例如薄膜 13 201226046 24之第一表面242的微粒或顆粒大小超過孔洞241尺寸的 固體物質，避免分離速度受到孔洞241上被截留物質堆積 的影響，造成操作阻力的增加。為產生較佳的平行剪切應 力及過濾效果，血液BL的流速範圍約由0.05 mL/min至 0.5 mL/min，其中，又以較高的流速可取得的血細胞分離 效果較好，例如以0.5 mL/min。 圖7係為應用本發明第一實施例之血液過濾方法之裝 置於運作時流道内的部分放大示意圖。請參考圖7所示， 在本實施例中，首先可提供一上述之裝置2，其具有一具 有複數孔洞241之薄膜24。接著，以注射針筒連結鐵氟龍 橋接頭，於'流人口輸入血液BL，且驅動血液BL…於流道 22内流動。由於流道22與薄膜24間的結構及設置關係， 血液BL於流通過流道22中至少有部分期間會接觸薄膜 24，且實質上與薄膜24相對平行移動。由於血液BL中例 如血漿SR等液態部分可通過薄膜24之孔洞141，而其餘 例如包括白血球WC、紅血球RC以及血小板PL等之血細 胞此類固態物質則會受到孔洞241的截留，故收集通過薄 膜24的血液BL部分，即血漿SR，便可完成血液BL之分 離作業。特別值得一提的是，在上述結構中，應用本發明 方法達成使血液BL在分離過程得以符合掃流原理，形成 平行剪切應力，故能有效降低濾餅的形成，防止發生阻塞 以及溶血現象。 圖8為應用本發明第一實施例之血液分離方法之裝置 之另一態樣的示意圖。請參考圖8所示，在本實施態樣中， 14 201226046 應用血液分離方法之襄置8 實施例所述相同，故以下僅就:未提及之二成二致與前述 態樣之裝置8更包括-,區動單元85，=本實施 液於流道82内流通。其中 — 货勖力驅動血 驅動皁兀8 5連接户卞β η201226046 VI. Description of the Invention: [Technical Field] The present invention relates to a blood separation method, and more particularly to a blood separation method using a sweeping principle. [Prior Art] The important work of oxygen in the body for oxygen and energy transmission is one of the indispensable elements for the normal operation of the living system. Based on this, various blood-centered application technologies have been developed in the field of clinical treatment and medical examination. It is hoped that the cells or substances contained in the blood can be obtained/analyzed correctly and quickly to further solve the discomfort caused by the pain and improve. The quality of human life. Under this concept, blood separation or filtration technology, which is the basis of many advanced applications, has been regarded as one of the key development projects. At present, it is known that techniques for separating blood cells and a group can be mainly classified into four categories. The first is a centrifugal technique in which the whole blood is placed in a test tube through the characteristics of a large specific gravity of the gray cells, and the centrifugal force generated by the high-speed rotation is used to achieve the goal of separating the blood cells from the plasma. However, since the density and deposition rate of some blood cells are very close to or even overlapping with the plasma components, the centrifugation technique does not provide the required purity for some specific applications. Moreover, such techniques typically consume a large amount of samples and drugs, and the processing takes a long time and the overall cost is high. The second principle is based on the dielectrophoretic force principle, combined with the micro-electromechanical process, providing a non-uniform alternating electric field with special instruments and equipment, and the separation between the two is achieved by the different conductivity and dielectric constant between the blood cells and the plasma. However, 201226046 This technology is still limited by the complexity of the steps and the expensive equipment required, and the application range is slightly narrow. The non-contact high-wavelength laser system has developed a new technology in recent years, which uses a light source in the opposite direction to form a stable energy trap to clamp tiny particles. Although, the current doctrine theory believes that the use of such a tool method for separating blood will have the advantages of non-contact, non-invasive, etc., so that non-contact high-wavelength lasers are very much expected in this field, but the actual It is still difficult to overcome the problem of high equipment cost in industrial application, and it is quite difficult to carry out the exhibition. The deep filtration system is more commonly used in the four techniques, and the blood is driven in the vertical direction through the multilayer film structure, so that the blood cells and the like - the solid matter is left on the surface of the film. However, the biggest problem with this technique is that the formation of the cake on the surface of the film 'especially with the increase in the amount of blood introduced, the thickness of the cake will inevitably increase correspondingly' so that every time it takes a period of time, it must be rinsed and removed, which is quite expensive. More manpower. In addition, the more important xin is 'deep filtration, which tends to exert greater pressure on blood cells in the blood, causing hemolysis in the filtration process, causing the separation operation to fail. In summary, it is obvious that the technique of separating blood cells to obtain plasma is inevitable that the sample consumption is large, the equipment is expensive, the application cost is not met, the operation time is long, and/or the process steps are cumbersome. Therefore, how to provide a blood sputum method 'It does not need the cost of hunting surface equipment, so it has better application flexibility' and it responds quickly, consumes less sample, and is more economical in operation. More importantly, it can effectively prevent filter cake formation and hemolysis. The phenomenon of 'genesis has become an important issue. 201226046 SUMMARY OF THE INVENTION In view of the above problems, an object of the present invention is to provide a blood separation method and a blood separation method for avoiding occlusion and hemolysis, which do not require high-cost equipment and instruments, and therefore have better application flexibility and reaction. Rapidly, the consumption of sample is small, the operation is more economical, and more importantly, it can effectively avoid the formation of filter cake and hemolysis. To achieve the above object, a ash separation method according to the present invention comprises the steps of: providing a film having a plurality of pores; driving a blood flow, and substantially parallel to the film when contacting the film; and collecting blood through the film The part of the hole. Among them, the part of the blood through the film hole is ϋ. In one embodiment of the invention, the blood separation method of the present invention further comprises the step of blocking blood cells in the blood by the membrane and passing the other portions of the blood through the pores of the membrane. In one embodiment of the invention, the blood system is continuously provided while driving through the continuous contact film. In an embodiment of the invention, the film has a first surface and a second surface. The first surface has a plurality of arcuate faces, the holes are disposed between the arcuate faces, and the second surface corresponds to the holes, and has a plurality of recessed regions. In an embodiment of the invention, the pores have a pore size ranging from 1 micron to 50 microns. In an embodiment of the invention, the aperture has a pore size in the range of 1 micron. In an embodiment of the invention, the material of the film comprises a metal and/or an alloy. In an embodiment of the invention, the material of the film is an alloy. In an embodiment of the invention, the blood system is driven by a drive unit. In its 201226046, the drive unit is a pump, aspirator or a combination thereof. In one embodiment of the invention, the blood system is driven by a micro-pump. To achieve the above object, a blood separation method for avoiding occlusion and hemolysis according to the present invention comprises the steps of: providing a film having a plurality of pores; driving a blood flow, and substantially parallel to the film when contacting the film; Collect blood through the holes in the membrane. Among them, the portion of the blood that passes through the pores of the film is plasma. In one embodiment of the invention, the blood separation method of the present invention for avoiding occlusion and hemolysis further comprises the step of blocking blood cells in the blood by the membrane and passing the other portions of the blood through the pores of the membrane. In an embodiment of the invention, the film has a first surface and a second surface. The first surface has a plurality of arcuate faces, the holes are disposed between the arcuate faces, and the second surface corresponds to the holes, and has a plurality of recessed regions. As described above, the blood separation method according to the present invention and the blood separation method for avoiding occlusion and hemolysis allow blood to flow through a film having a hole in a substantially parallel manner, and then pass through the size of the hole to restrict blood cells in the blood. It is trapped on one side of the film to be separated from a liquid component such as plasma. Among them, it is important that, according to the sweeping principle, since the blood system and the film move relatively in parallel, it is advantageous to form parallel shear stress, thus effectively reducing the problem of filter cake formation during blood separation, avoiding hole clogging and hemolysis, and affecting The success rate and efficiency of blood separation operations. Compared with the prior art, the blood separation method of the present invention and the blood separation method for avoiding occlusion and hemolysis do not require expensive equipment, and simply combine, for example, a flow path for liquid flow and a hole having a proper aperture 7 201226046 The film works, significantly reducing the threshold for use and is suitable for use in hospitals or research units at all levels. Furthermore, the method of the present invention has characteristics suitable for application to a biochip, and is not only convenient to carry and transport, but also capable of performing analysis and detection with a small amount of samples, and is economically advantageous in operation. More importantly, the side effects such as filter cake formation and hemolysis, which are common in the prior art, can be eliminated by the shear stress formed by the flow of blood, avoiding the problem of manual cleaning. [Embodiment] Hereinafter, a blood separation method according to a preferred embodiment of the present invention and a blood separation method for avoiding occlusion and hemolysis will be described with reference to the accompanying drawings, wherein the same elements will be described with the same reference numerals. BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 is a flow chart showing the steps of a blood separation method according to a first embodiment of the present invention. Referring to FIG. 1, in the embodiment, the blood separation method comprises the steps of: providing a film having a plurality of holes (S11); driving a blood flow, and substantially moving parallel to the film when contacting the film ( S13); and collecting a portion of the blood through the pore of the film (S15). Wherein, the portion of the blood that passes through the pores of the membrane is plasma. Here, the blood separation method of the present embodiment is for separating plasma and blood cells in blood, and the blood cell line is, for example, a collective term for red blood cells, various white blood cells, and/or blood platelets. In order to make the details of the steps of the embodiment more clear, the following will be taken as an example, and the structure and composition of the device will be clearly described. Based on this, how to implement the method of the present invention on the device will be explained. However, it is to be understood that the device in the following embodiments is merely for convenience of description and is not intended to limit the invention. Of course, other devices or systems that can be used in conjunction with the methods of the present invention can be used to implement the teachings of the present invention and are not limited to any particular configuration. Fig. 2 is a schematic view of an apparatus to which the blood separation method of the first embodiment of the present invention is applied, and Fig. 3 is an exploded perspective view of the apparatus shown in Fig. 2. Referring to Fig. 2 and Fig. 3 at the same time, in the present embodiment, the device 2 for applying the blood separation method may be a micro device such as a micro biochip, which is formed into a thin rectangular parallelepiped. In terms of the detailed structure, the apparatus 2 may include a covering casing 21, a first-class passage 22, a collecting tank 23, and a film 24 for filtration. The cover case 21 can further have an upper flow path portion 211 / and a lower collection portion 212. The upper flow path portion 211 and the lower collection portion 212 may be two bodies integrally formed or separated, and the sizes thereof may be the same or different. In the present embodiment, the upper flow path portion 211 and the lower collecting portion 212 are separately formed in two parts, but the length, the width, and the height are respectively 4 cm, 0 2.5 cm, and 0.5 cm. After the upper flow path portion 211 and the lower collecting portion 212 are formed, the coating casing 21 is formed by sealing and sealing. Of course, in addition to the above embodiments, the size of the two may be adjusted as needed for the application, and is not a limitation. The material of the cladding casing 21 and the upper flow channel portion 211 and the lower collecting portion 212 may be, for example, polymethylmethacrylate (PMMA), poly-dimethylsiloxane (PDMS), or ring. Epoxy, metal or glass. Of course, other polymer materials with better mechanical strength and high biocompatibility properties can also be used. In the present embodiment, the material of the cladding casing 21 and its upper flow channel portion 211 9 201226046 and the lower collecting portion 212 is exemplified by polydimethyl methoxy oxane (PDMS), which is not cytotoxic and is suitable as a living body. The material in contact with the biological sample. In addition, polydimethyloxane also has good light transmittance and is easy to observe and other characteristics. The flow path 22, the collecting groove 23, and the film 24 are housed in the covering case 21. The flow path 22 is disposed in the upper flow path portion 211, the collection groove 23 is disposed in the lower collection portion 212, and the thin portion 24 is interposed between the flow path 22 and the collection groove 23. The flow passage 22 is a continuous liquid passage, and the bottom side is open, and the other upper side and the left and right sides are closed on three sides. The flow path 22 is open to the outside, for example, in the present embodiment, a circular opening is formed on the surface 213 of the covering case 21. The openings of the flow passages 22 are respectively a liquid input inlet 221 and an output outlet 222, which can be respectively connected with a polytetrafluoroethene (PTFE) bridge joint, for example, to be connected with the injection syringe for blood input. Use while avoiding the occurrence of external leakage. The overall path of the flow channel 22 is not limited in shape. Here, the spiral or spiral shape is taken as an example, and a liquid path with a long distance can be formed within a certain area, which is beneficial to the blood separation operation. . However, in other embodiments, the flow channel 22 can also be in the form of a simple straight or serrated shape, depending on the blood separation requirement and the proportion of the components in the bloodstream, and can effectively separate the solid and liquid substances in the blood. In the present embodiment, the flow path 22 is provided in the flow path portion 211 above the cladding casing 21 by means of potting. In detail, a polybiguanide-based decane having a fluid colloid property may be used to cover the master mold, and after the colloid is solidified, the portion formed by the polybismuthyl decane is separated from the master mold to form a 201226046 setting. The flow path 22 has a flow path portion 211 over the cladding case 21. Fig. 4 is an enlarged schematic view showing the flow path shown in Fig. 3. Referring to Fig. 4, the flow path 22 is provided at the center of the upper flow path portion 211, and the entire flow path 22 (hereinafter referred to as a swept area) is spirally or spirally shaped. The diameter of the sweeping zone is about 0.8 cm. The channel width d2 of the flow channel 22 is about 50 micrometers, and the structural height of the channel is about 100 micrometers. Of course, the above dimensions are not the limiting conditions of the present invention, but can be adjusted according to practical applications. In other embodiments, the flow is The channel width d2 of the track 22 can be from about 50 to 500 microns, such as 100 or 200 microns, and the structural height of the channel 22 can be from about 10 to 500 micrometers, such as 20 or 200 microns. In addition, during the formation of the flow path 22, the flow path portion 211 on the cladding casing 21 is formed into an outwardly opening inlet port 221 and an outflow port 222 through the design of, for example, a mold, where both are elongated tubes. Body (not shown in Figure 4). Referring to FIG. 3, the collecting groove 23 of the embodiment may be disposed in the lower collecting portion 212 according to the same manner as described above, and the corresponding flow path 22 is disposed on the other side of the film 24. The collecting groove 23 may be a circular groove having a diameter of 0.8 cm and a structural height of 200 μm. Of course, the size of the collection trough 23 can also be adjusted, for example, by the amount of blood input. In other embodiments, the height of the structure can be between 10 and 500 microns, for example, the height of the structure can be 20 or 200 microns. In this embodiment, the centers of the sweeping zone, the collecting groove 23, and the film 24 may be substantially overlapped, for example, and the film 24 is interposed between the flow path 22 and the collecting groove 23, in other words, the film 24 is also sandwiched between Between the upper flow path portion 211 and the lower collecting portion 212. The size and configuration of the film 24 are not particularly limited, and s 11 201226046 is in principle based on the principle of cooperating with the sweeping zone. Figure 5a is a partially enlarged schematic view of the film of Figure 2, and Figure 5b is a schematic view of the film of Figure 5a taken along line B-B. Referring to Figures 5a and 5b at the same time, in the present embodiment, the film 24 may be a circular film having a diameter of 1 cm and having a plurality of holes 241 thereon. The film 24 has a first surface 242 and a second surface 243 which are respectively the upper surface and the lower surface of the film 24. In the present embodiment, the first surface 242 and the second surface 243 of the film 24 observed under the macroscopic view are both approximately flat surfaces without undulations (as shown in Fig. 3). However, referring to FIG. 5b, if further observed at a microscopic angle, the first surface 242 has a plurality of upwardly convex curved surfaces 244, and the holes 241 are disposed between the curved surfaces 244, and the second surface 243 corresponds to The hole 241 has a plurality of recessed regions 245. The above structure will be described in detail. Referring to FIG. 5a and FIG. 5b simultaneously, in the embodiment, the curved surface 244 can be, for example, a circular arc surface, and includes a top end and a bottom end, and the bottom end is disposed at a periphery of the hole 241. The recessed area 245 may be circular and have an area larger than the area of the hole 241. Further, the holes 241 are circular perforations that open the first surface 242 to the second surface 243 of the film 24, and may have a diameter of, for example, 1 μm. However, in other embodiments of the invention, the holes 241 may be of other shapes and/or sizes, for example from 1 to 50 microns, in principle depending on the effect desired for blood separation. For example, in the case of completely separating blood cells in the blood, the diameter of the holes 241 is preferably 1 μm. The film 24 may be of a metal and/or alloy material and may be manufactured through several steps including a lithography process and electroforming. 12 201226046 First, a substrate can be provided first, and then a photoresist layer is formed on one surface of the substrate. Subsequently, a plurality of photoresist patterns are formed on the photoresist layer and exposed on a part of the surface of the substrate. A photomask is disposed above the photoresist layer, wherein the photomask has a plurality of patterns corresponding to the photoresist patterns. Thereafter, exposure and development are performed to form a circular resist pattern which is not in the present embodiment. An alloy layer is formed on the photoresist pattern and the substrate by an electric prayer method. Finally, the alloy layer is separated from the photoresist pattern and the substrate to form a film 24 for separating blood that can be used in the method of the present embodiment. • Figure 6 is a schematic illustration of the device of Figure 2 at a section line A-A. Referring to FIG. 6, in the present embodiment, since the flow path 22 is substantially vertically stacked on the first surface 242 of the film 24, when the blood BL enters and flows through the flow path 22, there is bound to be Part of the film is in contact with the film 24 and flows over the film 24 in a manner substantially parallel to the film 24. The flow path 22 communicates with the collecting hole 23 through the hole 241 of the film 24 by the configuration in which the bottom side is open, so that when the blood BL flows through the flow path 22, it is subjected to gravity. The curved surface 244 of a surface 241 flows to the hole 241 (the figure is still flat due to the giant angle, and the detailed structure can be referred to FIG. 5b), so that the portion of the blood substance BL such as plasma is smoothly passed through the hole 241. As for the solid matter contained in the blood BL, for example, the particle or the particle size exceeding the size of the hole 241 is trapped by the hole 241 to achieve the effect of blood separation and filtration. It should be particularly noted that the structure of the apparatus 2 for applying the blood separation method of the present invention allows the blood BL to conform to the principle of sweep filtration to generate parallel shear stress, thereby sweeping blood cells accumulated in the filtration process, for example, film 13 201226046 24 The first surface 242 has a particle or particle size that exceeds the size of the solid material of the hole 241, preventing the separation speed from being affected by the accumulation of trapped material on the hole 241, resulting in an increase in operational resistance. In order to produce better parallel shear stress and filtration effect, the flow rate of blood BL ranges from about 0.05 mL/min to 0.5 mL/min, and the blood cell separation effect obtained at a higher flow rate is better, for example, 0.5. mL/min. Fig. 7 is a partially enlarged schematic view showing the flow of the blood filtration method to which the first embodiment of the present invention is applied, which is placed in the flow path during operation. Referring to Figure 7, in the present embodiment, a device 2 as described above is first provided having a film 24 having a plurality of holes 241. Next, the Teflon bridge joint is connected by a syringe, and blood BL is input to the flow population, and the blood BL is driven to flow in the flow path 22. Due to the structure and arrangement between the flow channel 22 and the membrane 24, the blood BL contacts the membrane 24 during at least a portion of the flow through the flow channel 22 and moves substantially parallel to the membrane 24. Since the liquid portion such as plasma SR in the blood BL can pass through the pore 141 of the membrane 24, the remaining solid matter such as blood cells including white blood cells WC, red blood cells RC, and platelets PL are trapped by the pores 241, and are collected through the membrane 24 The blood part of BL, the plasma SR, can complete the separation of blood BL. It is particularly worth mentioning that, in the above structure, the method of the present invention is applied to achieve the parallel flow of the blood BL in the separation process to form a parallel shear stress, thereby effectively reducing the formation of the filter cake and preventing the occurrence of clogging and hemolysis. . Fig. 8 is a view showing another aspect of the apparatus to which the blood separation method of the first embodiment of the present invention is applied. Please refer to FIG. 8 , in the present embodiment, 14 201226046 is the same as described in the embodiment of the blood separation method. Therefore, the following only refers to the device 8 which does not mention the above-mentioned aspect. Further, the --moving unit 85, = the present embodiment, flows through the flow path 82. Where — the cargo drive force drives the saponin 8 5 to connect the 卞β η
流入口 821，較佳地係透過鐵氣龍橋接㈣it心 連接，且設置於包覆殼體之上側。驅動單: 821 以微型注射幫浦’啟動後可持續將由外接式儲=如 未示）輸人裝置8之血液向前推動，且使其維持慣^圖 從而形成與薄膜84間的相對移動。依據此條件，動進。’ 元85當然可以為其他能提供動力的模組或元件，例如 吸器、手〜動操作注射針筒或其他相似裝置，亦或.於流人2 821及流出口 822形气氣體壓力差或真空狀態，使血液能 夠被驅動的模組或元#。 b 圖9為依據本發明第二實施例之血液分離方法的步驟 流程圖。請參考圖9所示，在本實施例中，血液分離方法 包括以下步驟：提供一具有複數孔洞之薄膜（S91);驅動 一血液流動，且於接觸薄膜時，實質上與薄膜相對平行移 動（S93);藉由薄膜阻擋血液内之血細胞，且血液之其他 部分通過薄膜之孔洞（S95);以及收集血液通過薄膜之孔 洞之部分（S97)。由於步驟S91、S93及S97係分別與本 發明第一實施例中之步驟Sll、S13及S15相同，故可參 考前述内容，以下將針對步驟S95及本發明之第一實施例 中未敍述的部分進一步說明。 同樣以配合第一實施例之裝置為例，在本實施例中， 丨 .S-i 15 201226046 由於流道係疊設於具有孔洞的薄膜，當血液在流道内流動 而與薄膜接觸時，理論上應受到重力作用而由薄膜的孔洞 穿過。然而，包括紅血球、白血球及/或血小板的企細胞因 為顆粒大於孔洞，不但無法穿過孔洞，還受到薄膜的阻擋 或截留。反觀血液中其他部分，尤其是液態物質部分，則 可以順利通過孔洞，因此達成分離金液成分的目的。在 此，要特別說明的是，透過調整薄膜上孔洞的孔徑大小可 以改變受到薄膜阻擋物質的組成，同時亦改變血液通過薄 膜孔洞部分的組成。舉例而言，在本發明另一實施例中， 若將孔洞放大，則薄膜僅阻擋血液中體積較大的白血球， 而通過薄膜的血液'部分則會含有體積相對較小的紅▲球 及血小板。由此可知，在保有避免濾餅形成特性的情況 下，本發明方法具有相當大的應用彈性。 本發明另揭露一種避免阻塞及溶血之血液分離方 法，其包括以下步驟：'提供一具有複數孔洞之薄膜；驅動 一血液流動，且於接觸薄膜時，實質上與薄膜相對平行移 動；以及收集血液通過薄膜之孔洞之部分。然而，此避免 阻塞及溶血之血液分離方法及其步驟的.相關細節與搭配 的實施裝置大致均與前述揭露之血液過濾方法相同，且已 詳述於上，於此不再贅述，惟再次特別強調的是，本發明 之避免阻塞及溶血之血液分離方法仍係基於血液在符合 掃流原理的情況下透過薄膜進行過濾分離，是以血液形成 的平行剪切應力能達到清除濾餅與避免溶血之功效。 為驗證本發明方法之可行，以下將舉人類血液為一實 16 201226046 驗例，並配合第一實碑例所述之裝置，說明實際應用本發 明方法分離血液中的血細胞以收集血漿。 實驗例1 利用本發明方法進行人類血液的分離 - 依據第一實施例揭示的技術内容製成裝置，在此，該 裝置為一種微型生物晶片，其中具有一具有孔洞之薄膜。 首先，以1 mL針筒量取人類全血血液0.5 mL。利用鐵氟 龍橋接頭連結注射針筒將全血血液注入微型生物晶片 • 中。調整微量注射幫浦，使流速維持在0,05 mL/min至0.5 mL/min的範圍内，驅動血液於流道内相對薄膜平行移動。 在此使用的數組流速設定可以為0.5 mL/min、0.3 - mL/min、0.1 mL/min、0·07 mL/min 及 0.05 mL/min。持續 輸入大約10分鐘内即可注射完畢。藉由光學顯微鏡觀察 過濾結果，如圖10所示，在10倍顯微鏡下可觀察到血液 流通流道C，而通過薄膜發生血漿與血細胞分離的情況（如 0 箭號D所指）。 綜上所述，因依據本發明之血液分離方法係使血液以 實質上平行的方式流過具有孔洞的薄膜，再透過孔洞的尺 寸限制，將血液中的血細胞截留於薄膜之一側，從而與例 如血漿等液態成分分離。其中，重要的是，依據掃流原理， 由於血液係與薄膜相對平行移動，故利於形成平行剪切應 力，如此，能有效降低血液過濾時濾餅形成的問題，避免 孔洞阻塞以及溶血現象，影響血液分離作業的成功率及效 17 201226046 與習知技術相較，本發明之血液分離方法不需藉助成 本昂貴的儀器設備，而簡單結合例如可供液體流動的流道 以及具有適當孔徑孔洞的薄膜即可運作，顯著地降低使用 門檻，適合推廣至各f級的醫療院所或研究單位使用。再 者，本發明血液分離方法具有適於應用在生物晶片的特 性，不僅便於攜帶、運送，且能以少量的樣本完成分析與 檢測，操作上較符合經濟效益。更重要的是，習知技術中 常見的濾餅形成及溶血現象等副作用均可透過血液在流 動時形成的剪切應力加以消除，避免時常需要人工清理的 問題。 拟上所述僅為舉_例性「而非為限制性者。任何未脫離 本發明之精神與範疇，而對其進行之等效修改或變更，均 應包括於後附之申請專利範圍中。 j · 【圖式簡單說明】 圖1為依據本發巧第一實施例之血液分離方法的步驟 流程圖； 圖2為應用本發明第一實施例之血液分離方法之裝置 的不意圖， 圖3為圖2所示之裝置的分解示意圖； 圖4為圖3所示之流道的放大示意圖； 圖5a為圖2所示之薄膜的部分放大示意圖； 圖5b為圖5a所示之薄膜於剖面線B-B位置的示意圖； 圖6為圖2所示之裝置於剖面線A-A位置的示意圖； 18 201226046 圖7係為應用本發明第一實施例之血液過濾方法之裝 置於運作時流道内的部分放大示意圖； 圖8為應用本發明第一實施例之血液分離方法之裝置 之另一態樣的示意圖； 圖9為依據本發明第二實施例之血液分離方法的步驟 流程圖；以及 圖10為本發明第一實驗例之結果於10倍顯微鏡下觀 察的照片圖。 【主要元件符號說明】 2、8 :裝置 21、 81 :包覆殼體 211 :上流道部 212 :下收集部 213 :表面 22、 82 :流道 221、 821 :流入口 222、 822 :流出口 23 :收集槽 24、84 :薄膜 241 :孔洞 242 :第一表面 243 :第二表面The inflow port 821 is preferably connected through the iron gas bridge (four) and is disposed on the upper side of the cladding casing. The drive list: 821 is activated by the micro-injection pump after the start-up, and the blood of the input device 8 can be pushed forward by the external storage device (not shown) and maintained to maintain the relative movement between the film 84 and the film 84. According to this condition, move forward. 'Yuan 85 can of course be used for other modules or components that can provide power, such as suction, hand-operated injection syringes or other similar devices, or in the flow of gas 2 821 and outlet 822 gas pressure difference or vacuum State, a module or element that enables blood to be driven. Figure 9 is a flow chart showing the steps of the blood separation method according to the second embodiment of the present invention. Referring to FIG. 9, in the embodiment, the blood separation method comprises the steps of: providing a film having a plurality of holes (S91); driving a blood flow, and substantially moving parallel to the film when contacting the film ( S93); blocking the blood cells in the blood by the film, and the other part of the blood passes through the pores of the film (S95); and collecting the blood through the pores of the film (S97). Since the steps S91, S93, and S97 are the same as the steps S11, S13, and S15 in the first embodiment of the present invention, reference may be made to the foregoing, and the following will be directed to the step S95 and the portion not described in the first embodiment of the present invention. Further explanation. Also taking the device of the first embodiment as an example, in the present embodiment, 丨.Si 15 201226046, because the flow channel is stacked on the film having the hole, when the blood flows in the flow path and comes into contact with the film, theoretically Passing through the holes of the film by gravity. However, cells including red blood cells, white blood cells, and/or platelets are not able to pass through the holes because they are larger than the pores, and are also blocked or trapped by the film. In contrast, other parts of the blood, especially the liquid substance, can pass through the holes smoothly, thus achieving the purpose of separating the components of the gold liquid. Here, it should be particularly noted that by adjusting the pore size of the pores in the film, the composition of the film-blocking substance can be changed, and the composition of the blood passing through the pore portion of the film is also changed. For example, in another embodiment of the present invention, if the hole is enlarged, the film only blocks the large white blood cells in the blood, and the blood portion of the film contains a relatively small volume of red ▲ balls and platelets. . From this, it is understood that the method of the present invention has considerable application flexibility while maintaining the characteristics of avoiding filter cake formation. The invention further discloses a blood separation method for avoiding occlusion and hemolysis, comprising the steps of: providing a film having a plurality of holes; driving a blood flow, and substantially parallel movement with the film when contacting the film; and collecting blood Pass the part of the hole in the film. However, the blood separation method and the steps thereof for avoiding occlusion and hemolysis are substantially the same as the blood filtration method disclosed above, and have been described in detail above, and will not be described again, but again It is emphasized that the blood separation method for avoiding occlusion and hemolysis of the present invention is still based on the filtration of blood through the membrane in accordance with the principle of sweeping, and the parallel shear stress formed by blood can achieve the removal of the filter cake and the avoidance of hemolysis. The effect. In order to verify the feasibility of the method of the present invention, the following is a description of the human blood as a method, and in conjunction with the apparatus described in the first embodiment, the method of the present invention is used to separate blood cells in blood to collect plasma. Experimental Example 1 Separation of Human Blood by the Method of the Present Invention - The apparatus was fabricated in accordance with the teachings of the first embodiment, where the apparatus is a micro biochip having a film having a hole. First, 0.5 mL of human whole blood was taken from a 1 mL syringe. Use a Teflon bridge connector to connect the syringe to inject whole blood into the micro biochip. The microinjection pump was adjusted to maintain the flow rate in the range of 0,05 mL/min to 0.5 mL/min, driving the blood to move parallel to the membrane in the flow channel. The array flow rate used here can be set to 0.5 mL/min, 0.3-mL/min, 0.1 mL/min, 0·07 mL/min, and 0.05 mL/min. The injection can be completed within about 10 minutes. The results of the filtration were observed by an optical microscope. As shown in Fig. 10, the blood flow path C was observed under a 10-fold microscope, and plasma and blood cells were separated by a film (as indicated by 0 arrow D). In summary, the blood separation method according to the present invention allows blood to flow through a membrane having pores in a substantially parallel manner, and then restricts the size of the pores, thereby trapping blood cells in the blood on one side of the membrane, thereby For example, liquid components such as plasma are separated. Among them, it is important that, according to the sweeping principle, since the blood system and the film move relatively in parallel, it is advantageous to form parallel shear stress. Thus, the problem of filter cake formation during blood filtration can be effectively reduced, and hole blocking and hemolysis can be avoided, and the influence is affected. Success rate and effect of blood separation operation 17 201226046 Compared with the prior art, the blood separation method of the present invention does not require expensive equipment, but simply combines, for example, a flow path for liquid flow and a film having a proper aperture. It can be operated, significantly reducing the threshold of use, and is suitable for use in medical institutions or research units of all grades. Furthermore, the blood separation method of the present invention has characteristics suitable for application to a biochip, and is not only convenient for carrying and transporting, but also capable of performing analysis and detection with a small amount of samples, and is economically advantageous in operation. More importantly, side effects such as filter cake formation and hemolysis, which are common in the prior art, can be eliminated by the shear stress formed by the blood flowing, avoiding the need for manual cleaning. It is intended that the following description be used as a limitation and not a limitation of the invention BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a flow chart showing the steps of the blood separation method according to the first embodiment of the present invention; FIG. 2 is a schematic view showing the apparatus for applying the blood separation method according to the first embodiment of the present invention. 3 is an exploded view of the apparatus shown in FIG. 2; FIG. 4 is an enlarged schematic view of the flow path shown in FIG. 3; FIG. 5a is a partially enlarged schematic view of the film shown in FIG. Figure 6 is a schematic view of the position of the device shown in Figure 2 at a section line AA; 18 201226046 Figure 7 is a partial enlargement of the flow channel in the operation of the device of the first embodiment of the present invention. Figure 8 is a schematic view showing another aspect of the apparatus for applying the blood separation method of the first embodiment of the present invention; Figure 9 is a flow chart showing the steps of the blood separation method according to the second embodiment of the present invention; hair Photograph of the results of the first experimental example observed under a 10x microscope. [Explanation of main component symbols] 2, 8: Apparatus 21, 81: Covered casing 211: Upper flow path portion 212: Lower collecting portion 213: Surface 22 82: flow paths 221, 821: inflow ports 222, 822: outflow ports 23: collecting grooves 24, 84: film 241: holes 242: first surface 243: second surface
244 弧面 -S 19 201226046 245 :凹陷區 85 :驅動單元 86 :鐵氟龍橋接頭 A-A、B-B :剖面線 BL :血液 C :流道 D :箭號 dl :直徑 d2 :寬度 PL :血小板 _RC :紅血球： SR :血漿 S11〜S15 、 S91〜S97 WC :白血球244 arc surface-S 19 201226046 245 : recessed area 85 : drive unit 86 : Teflon bridge joint AA, BB : section line BL : blood C : flow path D : arrow dl : diameter d2 : width PL : platelet _RC : red blood cells: SR: plasma S11~S15, S91~S97 WC: white blood cells