WO2021056952A1 - 一种白细胞提取装置 - Google Patents

一种白细胞提取装置 Download PDF

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Publication number
WO2021056952A1
WO2021056952A1 PCT/CN2020/075732 CN2020075732W WO2021056952A1 WO 2021056952 A1 WO2021056952 A1 WO 2021056952A1 CN 2020075732 W CN2020075732 W CN 2020075732W WO 2021056952 A1 WO2021056952 A1 WO 2021056952A1
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blood cell
channel
outlet
inlet
red blood
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PCT/CN2020/075732
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English (en)
French (fr)
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倪中华
项楠
易红
朱树
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东南大学
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
    • C12M47/04—Cell isolation or sorting

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  • the invention relates to a cell separation device, in particular to a white blood cell extraction device.
  • the white blood cells in human blood usually account for 0.1% of the total human blood cells, and the rich information it contains can be used for the diagnosis of many diseases. For example: coronary heart disease, cancer, etc.
  • the current mainstream method of white blood cell extraction is to first lyse red blood cells with red blood cell lysis solution, and then centrifuge for many times to remove the lysed red blood cells to leave white blood cells.
  • this method is time-consuming and labor-intensive, requiring manual operation by experienced operators, and the centrifugal force during centrifugation has been shown to be harmful to cell viability.
  • active microfluidic devices have achieved great development. They usually sort white blood cells according to the different dielectric properties and magnetic susceptibility of different cells. However, the blood processing speed of these methods is extremely low, usually only per minute. Process 1-10 microliters of blood. Leukocyte extraction devices based on inertial microfluidics usually dilute the blood 50-100 times in advance to extract leukocytes. Therefore, the pre-dilution operation increases the complexity of the operation and reduces the blood processing speed. Therefore, how to extract highly active leukocytes from undiluted blood automatically, at a high speed, and stably has great application value in the early diagnosis and prognostic evaluation of diseases.
  • the object of the present invention is to provide a white blood cell extraction device that can achieve high-speed extraction of white blood cells while removing red blood cells.
  • the leukocyte extraction device of the present invention includes five parts, from top to bottom, an upper cover plate, an extraction module, a storage module, a lysis module, and a lower cover plate.
  • the upper cover plate is provided with a white blood cell channel and a lysis plate.
  • the red blood cell channel and the physiological saline channel, the white blood cell channel and the lysed red blood cell channel are respectively provided with outlets, the physiological saline channel is provided with an inlet;
  • the extraction module is provided with a spiral extraction channel, a white blood cell channel and a lysed red blood cell channel
  • the storage module is provided with a reaction channel
  • the lysis module is provided with a mixing channel
  • the lower cover plate is provided with a blood channel and a red blood cell lysate channel
  • the blood channel and the red blood cell lysate channel are respectively provided with Inlet
  • the end of the blood channel and the end of the red blood cell lysate channel of the lower cover plate are respectively connected to the head end of the mixing channel of the lysis module at the same time, and the end of the mixing channel on the lysis module communicates with the head end of the reaction channel, so
  • the time for the mixed solution to pass through the reaction channel is 3-5 minutes, and the head end of the spiral extraction channel is in communication with the end of
  • Two outlets are provided, respectively communicating with the head end of the white blood cell channel and the head end of the lysed red blood cell channel of the extraction module.
  • the end of the white blood cell channel and the end of the lysed red blood cell channel of the extraction module are respectively connected with the white blood cell channel and the lysed red blood cell channel of the upper cover plate.
  • the lysed red blood cell channels are connected.
  • the white blood cell channel of the upper cover plate includes a white blood cell inlet, a white blood cell flow channel, and a white blood cell outlet
  • the lysed red blood cell channel includes a red blood cell lysis solution inlet, a red blood cell lysis solution flow channel and a red blood cell lysis solution outlet
  • the physiological saline channel It includes a normal saline inlet, a normal saline channel and a normal saline outlet.
  • the white blood cell channel end and the lysed red blood cell channel end of the extraction module are respectively connected to the white blood cell channel inlet and the lysed red blood cell channel inlet of the upper cover plate.
  • the head end of the spiral extraction channel is connected with the physiological saline outlet of the upper cover plate;
  • the reaction channel on the storage module includes a mixed liquid inlet, a mixed liquid flow channel and a mixed liquid outlet, and the mixed liquid outlet on the storage module is connected to the head end of the spiral extraction channel of the extraction module;
  • the mixing channel on the lysis module includes a red blood cell lysis solution inlet, a blood inlet, a mixing flow channel, and a mixed solution outlet, and the mixed solution outlet is connected to the mixed solution inlet of the storage module;
  • the blood channel on the lower cover plate includes a blood inlet, a blood flow channel, and a blood outlet
  • the red blood cell lysate channel includes a red blood cell lysis solution inlet, a red blood cell lysis solution flow channel, and a red blood cell lysis solution outlet.
  • the blood outlet of the lower cover plate and The red blood cell lysate outlet is respectively connected with the blood inlet and the red blood cell lysate inlet of the lysis module.
  • the blood outlet and the red blood cell lysate outlet of the lower cover plate are respectively provided with O-shaped gaskets, and the white blood cell inlet, the lysed red blood cell inlet, and the physiological saline outlet of the upper cover plate are respectively provided with O-shaped gaskets to facilitate communication with The connection of the corresponding interface.
  • the extraction module is formed by stacking a lower diversion layer, an extraction layer, and an upper diversion layer sequentially from bottom to top, the extraction layer is provided with one or more, and the lower diversion layer is provided with A mixed liquid inlet, a mixed liquid flow channel, and a mixed liquid outlet, the extraction layer is provided with a mixed liquid inlet, a physiological saline inlet, a spiral white blood cell extraction flow channel, a lysed red blood cell outlet, and a white blood cell outlet, the spiral white blood cell extraction flow channel
  • the end is provided with a Y-shaped double outlet, which is respectively connected with the lysed red blood cell outlet and the white blood cell outlet, and the upper guide layer is provided with a white blood cell inlet, a white blood cell flow channel, a white blood cell outlet, a lysed red blood cell inlet, and a lysed red blood cell flow channel.
  • Lysed red blood cell outlet physiological saline inlet, physiological saline channel, physiological saline outlet, white blood cell outlet of the upper diversion layer, lysed red blood cell outlet, saline inlet and white blood cell inlet of the upper cover, lysed red blood cell inlet, physiological
  • the saline outlet is connected, and the lysed red blood cell outlet, white blood cell outlet, and physiological saline inlet of the extraction layer are respectively connected with the lysed red blood cell inlet, white blood cell inlet, and physiological saline outlet of the upper diversion layer, and the mixed liquid outlet of the lower diversion layer is connected to the extraction layer
  • the mixed liquid inlet of the lower guide layer is connected with the mixed liquid outlet of the storage module.
  • each extraction layer is provided with one or more spiral-shaped leukocyte extraction flow channels, which are selected according to the amount of blood sample and the processing speed.
  • a double-sided adhesive layer is arranged between the layers of the extraction module for fixing, that is, the extraction module is composed of a lower diversion layer, a double-sided adhesive layer, an extraction layer two, a double-sided adhesive layer, an extraction layer one, and a double-sided adhesive layer.
  • the adhesive layer and the upper flow guiding layer are sequentially bonded and fixed from bottom to top, and the layers and layers of the double-sided adhesive layers are provided with escape holes at the junctions of the entrances and exits of the flow channels.
  • the storage module includes a chamber and a cover plate, the chamber is provided with at least two grooves, the cover plate is provided with at least one partition plate, and the partition plate is inserted into the groove
  • a mixed liquid flow channel is formed in the chamber, the first and last ends of the mixed liquid flow channel are respectively provided with a mixed liquid inlet and a mixed liquid outlet, and O-rings are respectively arranged at the mixed liquid inlet and the mixed liquid outlet .
  • the upper cover plate, the storage module and the lower cover plate are made of plastic or stainless steel
  • the O-shaped gasket is made of elastic materials such as rubber or silica gel
  • the extraction module and the pyrolysis module are made of polydimethylsiloxane.
  • PDMS photosensitive polymer
  • silica gel polydimethylsiloxane
  • the blood inlet is connected to a syringe containing blood
  • the red blood cell lysis solution inlet is connected to a syringe containing red blood cell lysis solution.
  • the blood and lysis solution pass through the blood inlet set on the lower cover plate and the red blood cell lysis solution respectively.
  • the inlet enters the blood flow channel and the red blood cell lysate flow channel, and is discharged from the blood outlet and red blood cell lysate outlet to the lysis module;
  • the blood and red blood cell lysate enter the lysis module through the blood inlet and the red blood cell lysate inlet respectively, and then the blood and lysate are mixed in the lysis module, and the final mixed solution is output to the storage module through the mixed solution outlet;
  • the mixed liquid enters the mixed liquid flow channel with a specific length path through the mixed liquid inlet of the storage module, and it takes 3-5 minutes for the mixed liquid to flow through the mixed liquid flow channel to the mixed liquid outlet to ensure the complete lysis of red blood cells;
  • the mixed solution of complete red blood cell lysis enters the mixed solution channel through the mixed solution inlet of the extraction module, and the physiological saline from the upper cover enters the physiological saline channel through the physiological saline inlet, and both enter the extraction layer at the same time.
  • the spiral extraction channel unit is used to extract white blood cells while removing lysed red blood cells; the principle is: when the fluid flows in the curved channel, the white blood cells and the lysed red blood cells are subjected to inertial lift and Dean's pulling force, and the white blood cells are balanced in the spiral On the inner wall of the flow channel, the lysed red blood cells circulate to the outer wall, and are finally collected and transported to the upper cover through the Y-shaped outlet;
  • the inlet of normal saline is connected with a syringe filled with normal saline.
  • the outlet of white blood cells and the outlet of lysed red blood cells are respectively connected to the white blood cell collection tube and the lysed red blood cell collection tube; the white blood cells and lysed red blood cells are finally led to the Different collection tubes.
  • the white blood cell extraction device related to the present invention can extract highly active white blood cells from undiluted blood automatically, at high speed, and stably, while removing lysed red blood cells. It provides a quick and effective way for subsequent clinical diagnosis of diseases.
  • Figure 1 is an exploded schematic diagram of the assembly of the leukocyte extraction device of the present invention
  • Figure 2 is a schematic diagram of the structure of the upper cover
  • Figure 3 is an exploded schematic diagram of the assembly of the extraction module
  • Figure 4 is a schematic diagram of the structure of the extraction module
  • Figure 5 is an exploded schematic diagram of the assembly of the storage module
  • Figure 6 is a schematic diagram of the structure of the storage module
  • Figure 7 is a schematic diagram of the structure of the cracking module
  • Figure 8 is a schematic view of the structure of the lower cover
  • Fig. 9 is the test results of various parameters of the present invention and the traditional white blood cell extraction method.
  • the white blood cell extraction device of the present invention is assembled from top to bottom by an upper cover 1, an extraction module 2, a storage module 3, a lysis module 4, and a lower cover 5.
  • the edge position of each part is uniform. Is provided with positioning installation holes;
  • the upper cover 1 is provided with a white blood cell outlet 11, a white blood cell flow channel 110, a white blood cell inlet 12, a lysed red blood cell outlet 13, a lysed red blood cell flow channel 19, a lysed red blood cell inlet 14, and physiological saline.
  • the extraction module 2 is composed of an upper diversion layer 21, a double-sided adhesive layer 22, an extraction layer 23, a double-sided adhesive layer 24, an extraction layer two 25, a double-sided adhesive layer 26, and a lower diversion layer.
  • Layer 27 is bonded and fixed sequentially from top to bottom;
  • the upper guide layer 21 is provided with a white blood cell outlet 211, four symmetrically arranged white blood cell inlets 212, a white blood cell flow channel 213, a lysed red blood cell inlet 214, a lysed red blood cell outlet 215, and a lysed red blood cell Flow channel 216, physiological saline inlet 217, four symmetrically arranged physiological saline outlets 218, and physiological saline flow channel 219;
  • the double-sided adhesive layer 22 is provided with through holes to facilitate the connection of the interface between two adjacent layers
  • the extraction layer 23 is provided with a mixed solution inlet 231, a physiological saline inlet 232, a spiral white blood cell extraction channel 233, a lysed red blood cell outlet 234, and a white blood cell outlet 235;
  • the double-sided adhesive layer 24 is provided with Through holes, it is convenient to connect the interface between two adjacent layers;
  • the extraction layer 25 is provided with a mixture inlet 251, a physiological saline inlet 252,
  • the spiral white blood cell extraction flow channels of the extraction layer one and the extraction layer two are arranged symmetrically along the X axis and the Y axis, respectively.
  • the double-sided adhesive layer 26 is provided with through holes to facilitate the connection of the interface between two adjacent layers;
  • the lower guide layer 27 is provided with a mixed liquid inlet 271 and four symmetrically arranged mixed liquid outlets 272.
  • the storage module 3 is provided with a chamber 31, a cover plate 32, an O-ring 33, a mixed liquid inlet 34, a mixed liquid outlet 35, and a mixed liquid flow channel 36.
  • the storage module 3 is formed by stacking a chamber 31 and a cover plate 32 from top to bottom.
  • the lysis module 4 is provided with a blood inlet 41, an red blood cell lysis solution inlet 42, and a mixed solution outlet 43.
  • the lower cover plate 5 is provided with a blood inlet 51, a blood outlet 52, a red blood cell lysis solution inlet 53, a red blood cell lysis solution outlet 54, an O-ring 55, a blood flow channel 56, and a red blood cell lysis solution flow. Road 57.
  • the present invention relates to a leukocyte extraction device.
  • the overall structure is composed of an upper cover plate 1, an extraction module 2, a storage module 3, a lysis module 4, and a lower cover plate 5, which are installed layer by layer from top to bottom.
  • the blood inlet 51 and the red blood cell lysate inlet 53 of the lower cover plate 5 are respectively connected to a syringe containing blood and red blood cell lysate, and are used for sampling the red blood cell lysate and physiological saline of the device; the blood outlet 52 and red blood cell
  • the lysis solution outlet 54 is respectively connected to the blood inlet 41 and the red blood cell lysis solution inlet 42 of the lysis module 4, and the lysis module 4 is used to thoroughly mix the red blood cell lysis solution and blood, and to achieve the preliminary lysis of the red blood cells;
  • the mixed solution outlet 43 of the module is connected to the mixed solution inlet 34 of the storage module.
  • the storage module 3 is provided with a long curved flow channel. It takes about 3-5 minutes to completely lyse the red blood cells in the blood. Therefore, from The mixed solution flowing out of the lysis module is diverted to the storage module. When the mixed solution flows to the outlet of the storage module, it takes 3-5 minutes to complete the complete lysis of red blood cells; the extraction module 2 integrates eight spiral inertias in parallel Sorting chips. Each sorting chip has two inlets and two outlets.
  • the mixed liquid outlet 35 of the storage module is connected to the mixed liquid inlet 271 of the lower guiding layer 27, and the mixed liquid outlet 271 of the lower guiding layer 27 is connected to the mixed liquid outlet 271 of the lower guiding layer 27.
  • the mixed liquid inlet 231 of the first extraction layer is connected with the mixed liquid inlet 251 of the second extraction layer at the same time.
  • the mixed liquid passes through the mixed liquid inlet to the lower diversion channel, and the lower diversion channel divides the mixed liquid evenly Up to eight spiral extraction units;
  • the saline outlet 16 of the upper cover plate is connected with the saline inlet 217 of the upper diversion layer, the saline outlet 218 is connected with the saline inlet 232 of the extraction layer 23, and at the same time with the extraction layer 25
  • the normal saline inlet 252 is connected, so the normal saline passes through the normal saline inlet 218 to the upper diversion channel, and the upper diversion channel is evenly divided into eight spiral extraction units.
  • the white blood cells and lysed cells obtained by the spiral extraction unit are processed.
  • the red blood cells are respectively guided to the white blood cell outlet 211 and the lysed red blood cell outlet 215 through the upper flow channel; as shown in FIG. 3, the spiral extraction unit is a micro-mixing microfluidic chip, and the height of the extraction flow channel is 80- 120 micrometers.
  • four sorting chips are integrated on the same plane and stacked in the vertical direction by a double-sided adhesive layer.
  • This embodiment relies on double The surface adhesive layer is stacked with an upper diversion layer and a lower diversion layer.
  • the white blood cell outlet 211 and the lysed red blood cell outlet 215 of the upper diversion layer are respectively connected to the white blood cell inlet 12 and the lysed red blood cell inlet 14 of the upper cover plate, and the physiological saline inlet 15 and the white blood cell outlet of the upper cover plate 1 11 and the lysed red blood cell outlet 13 are respectively connected with a syringe filled with physiological saline, a white blood cell collection tube, and a lysed red blood cell collection tube.
  • this embodiment can directly extract leukocytes from blood in about five minutes. As shown in Figure 9, the extraction efficiency can reach 83.9%, the cell activity rate is as high as 96.6%, and the red blood cell removal rate is as high as 98.2%.
  • the main materials of the upper cover plate 1 and the lower cover plate 5 are acrylic plates
  • the white blood cell outlet 11, the lysed red blood cell outlet 13, the physiological saline inlet 15, the blood inlet 51, and the red blood cell lysate inlet 53 are made of stainless steel
  • O type The sealing rings 17 and 55 are made of rubber.
  • the main body of the upper cover 1 and the lower cover 4 are machined from acrylic plates.
  • the white blood cell outlet 11, the lysed red blood cell outlet 13, the physiological saline inlet 15, the blood inlet 51, and the red blood cell lysate inlet 53 are made of stainless steel tubes. Use AB glue to bond with the machined acrylic board.
  • the upper diversion layer 21, the first diversion layer 23, the second diversion layer 25, and the lower diversion layer 27 in the extraction module 2 all have a three-layer structure.
  • the middle layer is made of silica gel, and the outer two layers are made of PVC plastic.
  • lasers are used to engrave the required structures on the selected PVC substrate and silica gel substrate respectively, and then the encapsulation is completed by ionic bonding technology.
  • the processing and manufacturing method of the cracking module 4 is similar to that of the extracting module 2. This technology has short processing time ( ⁇ 1min/piece), high processing accuracy (deviation of about 5 ⁇ m), low production cost and strong flexibility.

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Abstract

本发明公开了一种白细胞提取装置，自上而下依次包括上盖板、提取模块、存储模块、裂解模块和下盖板五部分，所述上盖板设置有白细胞通道、裂解的红细胞通道和生理盐水通道，所述白细胞通道和裂解的红细胞通道分别设有出口，所述生理盐水通道设有入口；所述提取模块上设置有螺旋形提取通道、白细胞通道和裂解的红细胞通道，所述的存储模块上设置有反应通道，所述裂解模块上设置有混合通道，所述下盖板上设置有血液通道和红细胞裂解液通道；本发明的白细胞提取装置，能够从未经稀释的血液中自动、高速、稳定地提取较高活性的白细胞，同时去除裂解的红细胞，为后续疾病的临床诊断等应用提供了一种快捷有效的方式。

Description

一种白细胞提取装置 技术领域
本发明涉及一种细胞分离装置，尤其涉及一种白细胞提取装置。
背景技术
人体血液中的白细胞通常占人体血细胞总量的0.1％，并且其含有的丰富信息可供许多疾病的诊断。例如：冠心病、癌症等。目前主流的白细胞提取方法是先用红细胞裂解液裂解红细胞，然后多次离心去除裂解的红细胞以留下白细胞。但是这种方法费时费力，需要有一定经验的操作人员手动操作，而且离心时的离心力已经被证明对细胞的活性有损害。
近年来，主动微流控装置获得了较大的发展，其通常根据不同细胞具有不同的介电特性、磁化率等来分选白细胞，但是这些方法的血液处理速度极低，通常每分钟只能处理1-10微升的血液。基于惯性微流控的白细胞提取装置通常要对血液提前进行50-100倍的稀释以提取白细胞，所以预稀释操作增加了操作的繁琐性、降低了血液处理速度。因此，如何能够从未经稀释的血液中自动、高速、稳定地提取较高活性的白细胞在疾病的早期诊断、预后评估等方面具有较大的应用价值。
发明内容
发明目的：本发明的目的是提供一种能够实现白细胞的高速提取，同时去除红细胞的白细胞提取装置。
技术方案：本发明所述的一种白细胞提取装置，自上而下依次包括上盖板、提取模块、存储模块、裂解模块和下盖板五部分，所述上盖板设置有白细胞通道、裂解的红细胞通道和生理盐水通道，所述白细胞通道和裂解的红细胞通道分别设有出口，所述生理盐水通道设有入口；所述提取模块上设置有螺旋形提取通道、白细胞通道和裂解的红细胞通道，所述的存储模块上设置有反应通道，所述裂解模块上设置有混合通道，所述下盖板上设置有血液通道和红细胞裂解液通道，所述血液通道和红细胞裂解液通道分别设有入口，所述下盖板的血液通道末端和红细胞裂解液通道末端分别同时与所述裂解模块的混合通道首端连通，所述裂解模块上的混合通道末端与所述反应通道首端连通，所述混合液通过反应通道的时间为3-5分钟，所述螺旋形提取通道首端同时与所述存储模块的反应通道末端和上盖板的生理盐水通道末端连通，所述螺旋形提取通道末端设有两个出口，分别与提取模块的白细胞通道首端和裂解的红细胞通道首端连通，所述提取模块的白细胞通道末端和裂解的红细胞通道的末端分别与所述上盖板的白细胞通道和裂解的红细胞通道连通。
优选的，所述上盖板的白细胞通道包括白细胞入口、白细胞流道和白细胞出口，所述裂解的红细胞通道包括红细胞裂解液入口、红细胞裂解液流道和红细胞裂解液出口，所述生理盐水通道包括生理盐水入口、生理盐水流道和生理盐水出口，所述提取模块的白细胞通道末端和裂解的红细胞通道的末端分别与所述上盖板的白细胞通道入口和裂解的红细胞通道入口 相连，所述螺旋形提取通道首端与所述上盖板的生理盐水出口相连；
所述存储模块上的反应通道包括混合液入口、混合液流道和混合液出口，所述存储模块上的混合液出口与所述提取模块的螺旋形提取通道首端相连；
所述裂解模块上的混合通道包括红细胞裂解液入口、血液入口、混合流道、混合液出口，所述混合液出口与所述存储模块的混合液入口相连；
所述下盖板上的血液通道包括血液入口、血液流道和血液出口，红细胞裂解液通道包括红细胞裂解液入口、红细胞裂解液流道和红细胞裂解液出口，所述下盖板的血液出口和红细胞裂解液出口分别与所述裂解模块的血液入口和红细胞裂解液入口相连。
优选的，所述下盖板的血液出口和红细胞裂解液出口分别设置有O型垫圈，所述上盖板的白细胞入口、裂解的红细胞入口、生理盐水出口上分别设置有O型垫圈，便于与相应接口的连接。
优选的，所述的提取模块由下导流层、提取层、上导流层自下而上依次堆叠而成，所述提取层设置有1或多个，所述下导流层上设置有混合液入口、混合液流道、混合液出口，所述提取层设置有混合液入口、生理盐水入口、螺旋形白细胞提取流道、裂解的红细胞出口、白细胞出口，所述螺旋形白细胞提取流道末端设有Y型双出口，分别与所述裂解的红细胞出口和白细胞出口相连，所述上导流层上设置有白细胞入口、白细胞流道、白细胞出口、裂解的红细胞入口、裂解的红细胞流道、裂解的红细胞出口、生理盐水入口、生理盐水流道、生理盐水出口，上导流层的白细胞出口、裂解的红细胞出口、生理盐水入口分别与上盖板的白细胞入口、裂解的红细胞入口、生理盐水出口相连，所述提取层的裂解的红细胞出口、白细胞出口、生理盐水入口分别与上导流层裂解的红细胞入口、白细胞入口、生理盐水出口相连，下导流层的混合液出口与提取层的混合液入口相连，所述下导流层的混合液入口和所述存储模块的混合液出口相连。
优选的，所述每个提取层上设置有1或者多个螺旋形白细胞提取流道，根据血样的量和处理速度进行选择。
优选的，所述提取模块的层与层间设置有双面胶层进行固定，即提取模块由下导流层、双面胶层、提取层二、双面胶层、提取层一、双面胶层和上导流层自下而上依次粘结固定，所述各双面胶层的层与层间在各流道出入口连接处设置有避让孔。
优选的，所述存储模块包括腔室和盖板，所述腔室中设置有至少两个凹槽，所述盖板上设置有至少1个隔板，所述隔板插入到所述凹槽中在腔室中形成混合液流道，所述混合液流道的首、末两端分别设置有混合液入口和混合液出口，所述混合液入口和混合液出口处分别设置有O型垫圈。
优选的，所述的上盖板、存储模块和下盖板为塑料或者不锈钢材质，所述的O型垫圈为橡胶或者硅胶材质等弹性材料，所述提取模块和裂解模块为聚二甲基硅氧烷PDMS、硅胶、塑料、玻璃材质中的一种或几种。
原理说明：
下盖板中，血液入口与装有血液的注射器相连，红细胞裂解液入口与装有红细胞裂解液的注射器相连，使用时，血液和裂解液分别通过设置在下盖板上的血液入口与红细胞裂解液入口进入血液流道和红细胞裂解液流道，并且由血液出口和红细胞裂解液出口排出至裂解模块；
裂解模块中，血液和红细胞裂解液分别通过血液入口、红细胞裂解液入口进入裂解模块，然后在裂解模块中完成血液和裂解液的混合，最终的混合液通过混合液出口输出至存储模块；
存储模块中，混合液通过存储模块的混合液入口进入具有特定长度路径的混合液流道，并且混合液经混合液流道流至混合液出口用时3-5分钟以保证红细胞的彻底裂解；
提取模块中，红细胞彻底裂解的混合液通过提取模块的混合液入口进入混合液流道，来自上盖板的生理盐水通过生理盐水入口进入到生理盐水流道，两者同时进入各提取层中设置的螺旋提取流道单元，以实现提取白细胞的同时去除裂解的红细胞；其原理是：流体在弯曲的流道中流动时，白细胞以及裂解的红细胞受到惯性升力以及Dean拽力的作用，白细胞平衡在螺旋流道的内壁面，裂解的红细胞循环至外壁面，最终通过Y型出口分别收集起来输送至上盖板；
上盖板中，生理盐水入口与装有生理盐水的注射器相连，白细胞出口、裂解的红细胞出口分别于白细胞收集管、裂解的红细胞收集管相连；白细胞和裂解的红细胞最终通过上盖板导流至不同的收集管中。
有益效果：本发明涉及的一种白细胞提取装置，能够从未经稀释的血液中自动、高速、稳定地提取较高活性的白细胞，同时去除裂解的红细胞。为后续疾病的临床诊断等应用提供了一种快捷有效的方式。
附图说明
图1是本发明白细胞提取装置的装配爆炸示意图；
图2是上盖板的结构示意图；
图3是提取模块的装配爆炸示意图；
图4是提取模块的结构示意图；
图5是存储模块的装配爆炸示意图；
图6是存储模块的结构示意图；
图7是裂解模块的结构示意图；
图8是下盖板的结构示意图；
图9是本发明和传统的白细胞提取方式的各个参数的测试结果。
具体实施方式
为进一步了解本发明的内容，结合附图及实施例对本发明作详细描述。
如图1所示，本发明所述白细胞提取装置由上盖板1、提取模块2、存储模块3、裂解模块4、下盖板5自上而下组装而成，每个部分的边缘位置均匀的设置有定位安装孔；
如图2所示，所述的上盖板1上设置有白细胞出口11、白细胞流道110、白细胞入口12、裂解的红细胞出口13、裂解的红细胞流道19、裂解的红细胞入口14、生理盐水入口15、生理盐水出口16、生理盐水流道18；其中，白细胞入口12、裂解的红细胞入口14、生理盐水出口16上设置有O型垫圈17；
如图3所示，所述的提取模块2由上导流层21、双面胶层22、提取层一23、双面胶层24、提取层二25、双面胶层26、下导流层27依次由上而下粘结固定；
如图4所示，所述的上导流层21上设置有白细胞出口211、四个对称布置的白细胞入口212、白细胞流道213、裂解的红细胞入口214、裂解的红细胞出口215、裂解的红细胞流道216、生理盐水入口217、四个对称布置的生理盐水出口218、生理盐水流道219；所述的双面胶层22上设置有通孔，便于相邻的两层之间接口的连接；所述的提取层一23上设置有混合液入口231、生理盐水入口232、螺旋形白细胞提取流道233、裂解的红细胞出口234、白细胞出口235；所述的双面胶层24上设置有通孔，便于相邻的两层之间接口的连接；所述的提取层二25上设置有混合液入口251、生理盐水入口252、螺旋形白细胞提取流道253、裂解的红细胞出口254、白细胞出口255；所述提取层一和提取层二的螺旋形白细胞提取流道分别沿X轴、Y轴对称布置4个。所述的双面胶层26上设置有通孔，便于相邻的两层之间接口的连接；所述的下导流层27上设置有混合液入口271、四个对称布置的混合液出口272、混合液流道273。
如图5所示，所述的存储模块3设置有腔室31、盖板32、O型垫圈33、混合液入口34、混合液出口35、混合液流道36。
如图6所示，所述的存储模块3由腔室31、盖板32自上而下堆叠而成。
如图7所示，所述的裂解模块4设置有血液入口41、红细胞裂解液入口42、混合液出口43。
如图8所示，所述的下盖板5上设置有血液入口51、血液出口52、红细胞裂解液入口53、红细胞裂解液出口54、O型垫圈55、血液流道56、红细胞裂解液流道57。
本发明涉及一种白细胞提取装置，整体结构由上自下依次由上盖板1、提取模块2、存储模块3、裂解模块4、下盖板5逐层安装而成。所述的下盖板5的血液入口51和红细胞裂解液入口53分别与装有血液和红细胞裂解液的注射器相连，用于本装置的红细胞裂解液和生理盐水的进样；血液出口52和红细胞裂解液出口54分别与所述裂解模块4的血液入口41、红细胞裂解液入口42相连，所述的裂解模块4用于红细胞裂解液和血液的彻底混合，并实现红细胞的初步裂解；所述裂解模块的混合液出口43和所述存储模块的混合液入口34相连，所述的存储模块3设置有弯曲的长流道，血液中的红细胞实现彻底的裂解约需3-5分钟，所以， 从裂解模块流出的混合液导流至存储模块，当混合液流至存储模块出口时耗时3-5分钟，正好完成红细胞的彻底裂解；所述的提取模块2并联集成了八个螺旋状的惯性分选芯片，每个分选芯片都有两个入口和两个出口，存储模块的混合液出口35和下导流层27的混合液入口271相连，下导流层27的混合液出口271和提取层一的混合液入口231相连，并同时与提取层二的混合液入口251相连，使用时，混合液通过混合液入口通至下导流流道，下导流流道将混合液均分至八个螺旋提取单元；上盖板的理盐水出口16和上导流层的生理盐水入口217相连，生理盐水出口218和提取层一23的生理盐水入口232相连，并同时与提取层二25的生理盐水入口252相连，所以，生理盐水通过生理盐水入口218通至上导流流道，并且被上导流流道均分至八个螺旋提取单元，经过螺旋提取单元处理得到的白细胞和裂解的红细胞通过上导流流道分别导流至白细胞出口211和裂解的红细胞出口215；如图3所示，所述螺旋提取单元为微混合微流控芯片，所述提取流道的高度为80-120微米，本实施例中在同一个平面集成了四个分选芯片并在垂直方向上依靠双面胶层进行堆叠，为了进样以及收集样本的方便，本实施例又在上下两侧依靠双面胶层堆叠了上导流层、下导流层。所述上导流层的白细胞出口211和裂解的红细胞出口215分别与所述上盖板的白细胞入口12和裂解的红细胞入口14相连，所述的上盖板1的生理盐水入口15、白细胞出口11和裂解的红细胞出口13分别与装有生理盐水的注射器、白细胞收集管、裂解的红细胞收集管相连。经过实验测试，本实施例能够在五分钟左右直接从血液中提取得到白细胞，如图9所示，提取效率可达83.9％，细胞的活性率高达96.6％，红细胞的去除率高达98.2％。两次离心提取白细胞一般耗时20分钟左右，虽然提取效率可达85.6％，但是活性率仅有86.3％，且一次离心的红细胞去除率为97.2％。可见本白细胞提取装置的综合性能超出常规离心法。
本实施例中，上盖板1和下盖板5主体材料为亚克力板，白细胞出口11、裂解的红细胞出口13、生理盐水入口15、血液入口51、红细胞裂解液入口53材料为不锈钢，O型密封圈17和55由橡胶制成。制作时，上盖板1以及下盖板4主体由亚克力板机加工而成，由不锈钢管制成的白细胞出口11、裂解的红细胞出口13、生理盐水入口15、血液入口51、红细胞裂解液入口53用AB胶与机加工后的亚克力板粘接。
本实施例中，提取模块2中的上导流层21、提取层一23、提取层二25、下导流层27均为三层结构。中间层材料均为硅胶，外侧两层材料均为PVC塑料。制作时，在选取的PVC基底和硅胶基底上分别用激光器刻出所需的结构，再通过离子体键合技术完成封装。裂解模块4的加工制作方式与提取模块2类似。本技术加工时间短(<1min/片)，加工精度高(偏差约5μm)，制作成本低，灵活性极强。

Claims (9)

  1. 一种白细胞提取装置，其特征在于，自上而下依次包括上盖板(1)、提取模块(2)、存储模块(3)、裂解模块(4)和下盖板(5)五部分，所述上盖板设置有白细胞通道、裂解的红细胞通道和生理盐水通道，所述白细胞通道和裂解的红细胞通道分别设有出口，所述生理盐水通道设有入口；所述提取模块设置有螺旋形提取通道、白细胞通道和裂解的红细胞通道，所述的存储模块上设置有反应通道，所述裂解模块上设置有混合通道，所述下盖板上设置有血液通道和红细胞裂解液通道，所述血液通道和红细胞裂解液通道分别设有入口，所述下盖板的血液通道末端和红细胞裂解液通道末端分别同时与所述裂解模块的混合通道首端连通，所述裂解模块上的混合通道末端与所述反应通道首端连通，所述混合液通过反应通道的时间为3-5分钟，所述螺旋形提取通道首端同时与所述存储模块的反应通道末端和上盖板的生理盐水通道末端连通，所述螺旋形提取通道末端设有两个出口，分别与提取模块的白细胞通道首端和裂解的红细胞通道首端连通，所述提取模块的白细胞通道末端和裂解的红细胞通道的末端分别与所述上盖板的白细胞通道和裂解的红细胞通道连通。
  2. 根据根据权利要求1所述的一种白细胞提取装置，其特征在于，
    所述上盖板的白细胞通道包括白细胞入口(12)、白细胞流道(110)和白细胞出口(11)，所述裂解的红细胞通道包括红细胞裂解液入口(14)、红细胞裂解液流道(19)和红细胞裂解液出口(13)，所述生理盐水通道包括生理盐水入口(15)、生理盐水流道(18)和生理盐水出口(16)，所述提取模块的白细胞通道末端和裂解的红细胞通道末端分别与所述上盖板的白细胞入口和裂解的红细胞入口相连，所述螺旋形提取通道首端与所述上盖板的生理盐水出口(16)相连；
    所述存储模块上的反应通道包括混合液入口(34)、混合液流道(36)和混合液出口(35)，所述存储模块上的混合液出口(35)与所述提取模块的螺旋形提取通道首端相连；
    所述裂解模块上的混合通道包括红细胞裂解液入口(42)、血液入口(41)、混合流道、混合液出口(43)，所述混合液出口(43)与所述存储模块的混合液入口(34)相连；
    所述的下盖板上的血液通道包括血液入口(51)、血液流道(56)和血液出口(57)，红细胞裂解液通道包括红细胞裂解液入口(53)、红细胞裂解液流道(57)和红细胞裂解液出口(54)，所述下盖板的血液出口和红细胞裂解液出口分别与所述裂解模块的血液入口和红细胞裂解液入口相连。
  3. 根据权利要求2所述的一种白细胞提取装置，其特征在于，所述下盖板的血液出口和红细胞裂解液出口位置分别设置有O型垫圈(55)、所述上盖板的白细胞入口、裂解的红细胞入口、生理盐水出口位置分别设置有O型垫圈(17)。
  4. 根据权利要求2所述的一种白细胞提取装置，其特征在于，所述的提取模块由下导流层(27)、提取层(23)、上导流层(21)自下而上依次堆叠而成，所述提取层设置有1或多个，所述下导流层上设置有混合液入口(271)、混合液流道(273)、混合液出口(272)，所述提取层设置有混合液入口(231)、生理盐水入口(232)、螺旋形白细胞提取流道(233)、 裂解的红细胞出口(234)、白细胞出口(235)，所述螺旋形白细胞提取流道末端设有Y型双出口，分别与所述裂解的红细胞出口(234)和白细胞出口相连，所述上导流层上设置有白细胞入口(212)、白细胞流道(213)、白细胞出口(211)、裂解的红细胞入口(214)、裂解的红细胞流道(216)、裂解的红细胞出口(215)、生理盐水入口(217)、生理盐水流道(219)、生理盐水出口(218)，上导流层的白细胞出口、裂解的红细胞出口、生理盐水入口分别与上盖板的白细胞入口、裂解的红细胞入口、生理盐水出口相连，所述提取层的裂解的红细胞出口、白细胞出口、生理盐水入口分别与上导流层裂解的红细胞入口、白细胞入口、生理盐水出口相连，所述下导流层的混合液出口(273)与提取层一的混合液入口(231)相连，所述下导流层的混合液入口(271)和所述存储模块的混合液出口(35)相连。
  5. 根据权利要求4所述的一种白细胞提取装置，其特征在于，所述每个提取层上设置有1或者多个螺旋形白细胞提取流道。
  6. 根据权利要求4所述的一种白细胞提取装置，其特征在于，所述提取模块的层与层间设置有双面胶层进行固定，即提取模块由下导流层(27)、双面胶层(26)、提取层二(25)、双面胶层(24)、提取层一(23)、双面胶层(22)和上导流层(21)自下而上依次粘结固定，所述各双面胶层的层与层间在各流道出入口连接处设置有避让孔。
  7. 根据权利要求1所述的一种白细胞提取装置，其特征在于，所述存储模块包括腔室(31)和盖板(32)，所述腔室中设置有1或多个凹槽，所述盖板上设置有1或多个隔板，所述隔板插入到所述凹槽中在腔室中形成混合液流道(36)，所述混合液流道的首、末两端分别设置有混合液入口(34)和混合液出口(35)，所述混合液入口和混合液出口处分别设置有O型垫圈(33)。
  8. 根据权利要求1所述的一种白细胞提取装置，其特征在于，所述的上盖板、存储模块和下盖板为塑料或者不锈钢材质，所述的O型垫圈为橡胶或者硅胶材质，所述提取模块和裂解模块为聚二甲基硅氧烷PDMS、硅胶、塑料、玻璃材质中的一种或几种。
  9. 根据权利要求1所述的一种白细胞提取装置，其特征在于，所述上盖板、提取模块、存储模块、裂解模块和下盖板的边缘分别设有定位安装孔。
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