WO2022247246A1 - 用于质谱仪的上样装置 - Google Patents

用于质谱仪的上样装置 Download PDF

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Publication number
WO2022247246A1
WO2022247246A1 PCT/CN2021/139303 CN2021139303W WO2022247246A1 WO 2022247246 A1 WO2022247246 A1 WO 2022247246A1 CN 2021139303 W CN2021139303 W CN 2021139303W WO 2022247246 A1 WO2022247246 A1 WO 2022247246A1
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sample
mass spectrometer
loading device
microfluidic chip
sample loading
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French (fr)
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门涌帆
王冠博
李雯
柴胡玲潇
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Shenzhen Institute of Advanced Technology of CAS
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Shenzhen Institute of Advanced Technology of CAS
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    • H—ELECTRICITY
    • H01—ELECTRIC ELEMENTS
    • H01J—ELECTRIC DISCHARGE TUBES OR DISCHARGE LAMPS
    • H01J49/00—Particle spectrometers or separator tubes
    • H01J49/02—Details
    • H01J49/04—Arrangements for introducing or extracting samples to be analysed, e.g. vacuum locks; Arrangements for external adjustment of electron- or ion-optical components
    • H01J49/0431—Arrangements for introducing or extracting samples to be analysed, e.g. vacuum locks; Arrangements for external adjustment of electron- or ion-optical components for liquid samples
    • H01J49/0445—Arrangements for introducing or extracting samples to be analysed, e.g. vacuum locks; Arrangements for external adjustment of electron- or ion-optical components for liquid samples with means for introducing as a spray, a jet or an aerosol

Definitions

  • the invention relates to the technical field of biomedicine, in particular to a sample loading device for a mass spectrometer.
  • Mass spectrometry is a powerful method for targeting the structural and dynamic properties of native and unnatural proteins. Due to its high analytical sensitivity and specificity, MS has significant advantages in molecular and pharmacological analysis as well as in clinical practice. However, this technology still needs to improve performance in the following areas: (1) Dynamic (time) resolution capability. The high-order structure and interaction of proteins in solution environment are highly dynamic, and species can be rapidly interconverted, and a large number of intermediates can also be formed. However, existing methods are difficult to provide characterization data in the time dimension. (2) Operability. Some methods have many steps and require high experience and technical requirements for operators, so they still need to be simplified for large-scale promotion in industrial practice. (3) Integration.
  • Microfluidic devices offer various advantages in manipulating and detecting chemicals and biomolecules.
  • the platform can achieve low sample consumption, automation, high-throughput operation and temperature control, flexible parameter customization and adjustable reaction time.
  • a mass spectrometer sample loading device based on a microfluidic chip, samples can be completed online.
  • the pretreatment and biochemical reaction chip are operated, and samples are injected downstream in real time, so that the product characteristics under specific reaction time conditions can be accurately detected, and a lot of sample waste and labor costs are saved. Therefore, the combination of microfluidic chips and mass spectrometers is very needs.
  • analyte ionization is very important for MS sample analysis.
  • the fabrication and reproducibility of glass or polymer chip-based interfaces is not easy, which also limits the widespread use of online electrospray mass spectrometry (ESI-MS).
  • the object of the present invention is to provide a sample loading device for mass spectrometer, the device scheme can be adapted to the mass spectrometer on the total flow rate and solution composition after the sample is pretreated or reacted, and provides Stable and high ionization efficiency, and the chip-mass spectrometer interface design is convenient, simple and scalable.
  • the technical solution provided by the present invention is: a sample loading device for a mass spectrometer, the sample loading device is arranged on the side of the sample inlet of the mass spectrometer, and the sample loading device includes a microfluidic chip , connecting pipelines, injection needles and sample injection structures;
  • the sample injection structure injects the sample solution to be tested into the microfluidic chip
  • One end of the connecting pipeline is connected to the outlet port of the microfluidic chip
  • the spray needle is connected to the other end of the connecting pipeline, the spray needle is powered on, and the sample solution in the spray needle is sprayed by means of electrospray;
  • the needle generates a spray spray into the injection port of the mass spectrometer.
  • the connecting pipeline includes a two-way joint and a sleeve, one end of the two-way joint is connected to the outlet end of the microfluidic chip through the sleeve, and the other end of the two-way joint is connected through the sleeve A tube is connected to the needle.
  • the material of the sleeve is a polymer material, and the sleeve is interference-fitted with the outlet end of the microfluidic chip.
  • the included angle between the microfluidic chip and the spray needle is an acute angle, a right angle or an obtuse angle, and the connecting pipeline and the spray needle are on the same straight line.
  • the distance from the outlet of the needle to the inlet of the mass spectrometer is 1-100 mm.
  • the spray needle mentioned above includes but is not limited to a microporous sleeve-like structure made of stainless steel.
  • the spray needle can be simplified as a conductive capillary directly connected to the outlet of the microfluidic chip, without the need for two-way joints, sleeves and other parts.
  • the mass spectrometer includes a power supply, the needle is connected to the power supply of the mass spectrometer, and the mass spectrometer provides electric energy.
  • the inner diameter of the spray needle is 1-200 ⁇ m
  • the outer diameter is 100-3000 ⁇ m
  • the length is 1-100 mm.
  • the microfluidic chip includes a main channel and an auxiliary channel, and the main channel and the auxiliary channel are used to add different solutions;
  • the auxiliary flow channel is connected to the main flow channel, and there are multiple auxiliary flow channels.
  • a diversion structure is provided in the main channel, and the diversion structure is used for mixing different solutions added in the main channel and the auxiliary channel.
  • the flow guide structure includes a fishbone structure
  • the fishbone structure includes a first fishbone structure and a second fishbone structure
  • the first fishbone structure and the second fishbone structure are axisymmetric structures, so The first fishbone structure and the second fishbone structure are arranged at intervals in the main channel.
  • a connecting pipeline is used to connect the spray needle to the outlet end of the microfluidic chip, and then the sample solution is sprayed into the sample inlet of the mass spectrometer by electrospray technology.
  • Such arrangement can improve the electrospray efficiency and the efficiency of the mass spectrometer on the one hand.
  • Fig. 1 is a schematic structural view of a sample loading device and a mass spectrometer according to an embodiment of the present invention
  • Fig. 2 is a structural schematic diagram of a connecting pipeline and an injection needle according to an embodiment of the present invention
  • Fig. 3 is a sectional view of part A in Fig. 1;
  • FIG. 4 is a schematic structural view of a microfluidic chip according to an embodiment of the present invention.
  • Fig. 5 is a schematic diagram of the structure and solution mixing of the confluence of the main channel and the auxiliary channel according to an embodiment of the present invention
  • Fig. 6 is a schematic structural diagram of a microfluidic chip according to another embodiment of the present invention.
  • sample loading device 110, microfluidic chip; 111, main flow channel; 112, auxiliary flow channel; 113, fishbone structure; 113a, first fishbone structure; 113b, second fishbone structure; 120, connection 121, two-way joint; 122, casing; 130, injection needle; 140, sample injection structure; 141, sample loading pipeline; 142, solution injector; 200, mass spectrometer; 210, sample inlet; 220, power supply.
  • first and second are used for descriptive purposes only, and cannot be understood as indicating relative importance, or implicitly indicating the quantity of indicated technical features. Therefore, unless otherwise specified, the features defined as “first” and “second” may explicitly or implicitly include one or more of these features; “plurality” means two or more.
  • the term “comprising” and any variations thereof mean non-exclusive inclusion, possible presence or addition of one or more other features, integers, steps, operations, units, components and/or combinations thereof.
  • the terms “mounted”, “connected” and “connected” should be interpreted in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection , can also be an electrical connection; it can be a direct connection, an indirect connection through an intermediary, or an internal communication between two components.
  • the terms “mounted”, “connected” and “connected” should be interpreted in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection , can also be an electrical connection; it can be a direct connection, an indirect connection through an intermediary, or an internal communication between two components.
  • the embodiment of the present invention discloses a sample loading device 100 for a mass spectrometer 200, the sample loading device 100 is arranged on the side of the sample inlet 210 of the mass spectrometer 200, the sample loading device 100 includes a microfluidic chip 110, a connecting pipeline 120, a needle 130, and a sample injection structure 140; the sample injection structure 140 injects the sample solution to be tested into the microfluidic chip 110; one end of the connecting pipeline 120 is connected to At the outlet end of the microfluidic chip 110; the spray needle 130 is connected to the other end of the connecting pipeline 120, the spray needle 130 is connected to the power supply 220, and the spray needle 130 is made to The sample solution in the sample solution forms a spray; the spray needle 130 generates a spray and sprays it into the sample inlet 210 of the mass spectrometer 200 .
  • the principle of spraying the sample solution in the spray needle 130 by means of electrospray is: when the fine mist droplets are sprayed out from the metal spray needle 130, they are obtained from the high-intensity electric field at the mouth of the metal spray needle 130. A large number of charges, due to the Coulomb force between the charges, these charges are evenly distributed on the surface of the droplet. When the droplet is dried to remove the solvent, the volume of the droplet decreases gradually, so the charge per unit surface area increases sharply, making the droplet unstable and splitting, resulting in a finer droplet spray.
  • the nozzle 130 is connected to the outlet port of the microfluidic chip 110 by using a connecting pipeline 120, and then the sample solution is sprayed into the sample inlet 210 of the mass spectrometer 200 by electrospray technology.
  • the electrospray efficiency and sensitivity of the mass spectrometer 200 improve the identification of analytes with diverse structures;
  • the volume is small, and it is easy to replace, which simplifies the operation of replacing the microfluidic chip 110 or the injection needle 130 in the experiment, and reduces the situation that the experiment cannot be continued when the pollution or the injection needle 130 is blocked.
  • the mass spectrometer 200 includes a power supply 220 , the needle 130 is connected to the power supply 220 of the mass spectrometer 200 , and the mass spectrometer 200 provides electricity.
  • the sample loading device 100 is powered by the mass spectrometer 200 to perform electrospray without additional power supply 220 , which reduces the volume of the sample loading device 100 and increases the portability and ease of use of the sample loading device 100 .
  • the distance from the outlet of the injection needle 130 to the sample inlet 210 of the mass spectrometer 200 is 1-100 mm. Ensure that the tiny droplets formed by the electrospray of the sample solution can evenly fly to the sample inlet 210 of the mass spectrometer 200, and prevent the tiny droplets sprayed into the sample inlet 210 of the mass spectrometer 200 from being uneven, Or the distance is too long so that some formed tiny droplets cannot fly into the sample inlet 210 of the mass spectrometer 200 and fall outside to cause waste or pollution.
  • the connecting pipeline 120 includes a two-way joint 121 and a sleeve 122, and one end of the two-way joint 121 is connected to the microfluidic chip 110 through the sleeve 122.
  • the outlet end of the two-way joint 121 is connected to the spray needle 130 through the sleeve 122 .
  • the sleeve 122 is made of a polymer material, and the sleeve 122 is directly inserted into the outlet end of the microfluidic chip 110 through frictional force, and has an interference fit with the outlet end of the microfluidic chip 110, so that the connection is tight and leak-free.
  • the material of the sleeve 122 can be PEEK polyether ether ketone, PEK polyether ketone, PEKK polyether ketone ketone, PEEKK polyether ether ketone ketone, PEKEKK polyether ketone ether ketone ketone, PFA perfluoroalkoxy Resin, FEP fluorinated ethylene propylene, ETFE ethylene-tetrafluoroethylene or PTFE polytetrafluoroethylene and other polymer materials suitable for mass spectrometry experiments.
  • the spray needle 130 is a metal spray needle 130
  • the inner diameter of the spray needle 130 is 1-200 ⁇ m
  • the outer diameter is 100-3000 ⁇ m
  • the length is 1-100 mm
  • the inner diameter of the sleeve 122 is 150-200 ⁇ m.
  • the inner diameter of the spray needle 130 is 30 ⁇ m
  • the outer diameter is 150 ⁇ m
  • the length is 40 ⁇ m
  • the inner diameter of the sleeve 122 is 180 ⁇ m.
  • the needle 130 , the sleeve 122 and the two-way connector 121 are on the same straight line after being connected, and the included angle with the overall flow direction of the microfluidic chip 110 is an acute angle, a right angle or an obtuse angle.
  • the current connection method is to make the overall direction of the flow of the added sample solution be a straight line, which requires a more precise design, and the seal also requires a tight connection design. In this case, replacement is very troublesome, and precise Adjustment takes a long time, which will delay the detection of the sample solution or the progress of the experiment.
  • the microfluidic chip 110 includes a primary channel 111 and an auxiliary channel 112, and the sample injection structure 140 includes a sample loading pipeline 141 and a solution injector 142, and the upper
  • the sample pipeline 141 is respectively inserted into the inlets of the main flow channel 111 and the auxiliary flow channel 112, and the other end of the sample loading pipeline 141 is connected to the solution injector 142, and the sample solution and the added solution are respectively injected into the Among the main channel 111 and the auxiliary channel 112 ;
  • the auxiliary channel 112 is connected to the main channel 111 , there are multiple auxiliary channels 112 , and the interval between each auxiliary channel 112 is 11500 ⁇ m.
  • the microfluidic chip 110 can realize the on-line mixing of multiple liquids.
  • the main sample solution is added to the main channel 111, and the required additional liquid is added to the auxiliary channel 112 to complete the mixing in the subsequent channel, that is, in the microfluidic
  • the mixing and reaction of the solution can be completed in the chip 110, and the continuous flow operation can be used for mass spectrometry sample loading analysis. It is not necessary to mix the solution first and then use it for detection. To change the mixing ratio, the detection operation needs to be stopped. And by adjusting the injection pressure through the solution injector 142 to adjust the flow rate of the solution, different mixing ratios can be changed in real time to achieve different reaction mixing ratios, reaction times and mixing effects.
  • the protein hydrogen-deuterium exchange reaction time can be adjusted online in real time, realizing the operation of online dynamic analysis of protein structure.
  • Figure 5 it shows the mixing balance effect of the solution in the main channel 111 (CH1) and the solution in the auxiliary channel 112 (CH2).
  • the microfluidic chip 110 can achieve different ratios from 99:1 to 50:50. the mixing function.
  • the main channel 111 is provided with a guide structure, and the guide structure is used for mixing different solutions added in the main channel 111 and the auxiliary channel 112 .
  • the diversion structure is a herringbone structure 113 (Herringbone structure).
  • the herringbone structure 113 is arranged after the main channel 111 and the first auxiliary channel 112 merge. Location.
  • a vortex or chaotic flow will be formed, which can quickly mix uniformly, improve the efficiency of uniform mixing, and shorten the length of the flow channel required for mixing, so that the size of the microfluidic chip 110 can be reduced even more. for small.
  • the flow guide structure is not limited to the fishbone structure 113, and other geometric structures for mixing are also possible.
  • the fishbone structure 113 includes a first fishbone structure 113a and a second fishbone structure 113b, the first fishbone structure 113a and the second fishbone structure 113b are axisymmetric structures, and the first fishbone structure The structure 113a and the second fishbone structure 113b are arranged at intervals in the main channel 111 . Circular mixing is performed along the shapes of the first fishbone structure 113a and the second fishbone structure 113b, so that the mixing of the solution is more uniform and faster.
  • the microfluidic chip 110 includes a main flow channel 111 and two auxiliary flow channels 112, the main flow channel 111 is a main line flow channel, and the two auxiliary flow channels 112 are respectively connected to the main flow channel 111 Among them, the interval between the two auxiliary channels 112 is 4000 ⁇ m-11500 ⁇ m, depending on the number of cycles.
  • a first fishbone structure 113a plus a second fishbone structure 113b is a cycle, and eight cycles are set in the main channel 111, that is, eight first fishbone structures 113a and eight second fishbone structures 113b are spaced apart
  • the setting, that is, the interval between the two auxiliary flow channels 112 is set to 11500 ⁇ m.
  • the arrangement of the auxiliary flow channels 112 is not limited to the arrangement at intervals, and the arrangement of the auxiliary flow channels 112 may also be arranged in layers, or arranged at an included angle.
  • mass spectrometry elucidates protein structure.
  • the microfluidic chip 110 can realize the online mixing of multi-channel liquids.
  • the main sample solution is added to the main channel 111, and the required additional liquid is added to the auxiliary channel 112, and a circulating first herringbone structure 113a is added.
  • the second fishbone structure 113b quickly and uniformly mixes the solution and reacts the sample online, and performs continuous flow operation for mass spectrometry sample loading analysis.
  • one main flow channel 111 and two auxiliary flow channels 112 form a set of flow channels
  • the microfluidic chip 110 includes two independent sets of flow channels.
  • multiple flow channels can also be set.
  • the group of flow channels can be used multiple times without cleaning, and different sample solutions can be tested at the same time. It is relatively easy to directly design multiple sets of flow channels during the manufacture of the microfluidic chip 110 , and has little influence on the manufacturing difficulty, which can save raw materials and time.
  • the material of the microfluidic chip can be glass quartz, silicon wafer, paper-based material and high molecular polymer (PDMS, PMMA and PC, etc.).
  • the manufacturing method of the above-mentioned microfluidic chip 110 specifically includes: designing the microfluidic chip 110 with the fishbone structure 113 using graphics software, and making a film, and then making a microfluidic chip template by photolithography.
  • the exemplary microfluidic chip 110 is mixed with Polydimethylsiloxane (Polydimethylsiloxane, PDMS) A liquid and B liquid at a ratio of 10:1, and then poured into the photolithographic template prepared above, after vacuum degassing , place it horizontally in an oven at 80 degrees, let it stand for 45 minutes, and remove the mold to obtain the corresponding PDMS chip.
  • PDMS Polydimethylsiloxane
  • the microfluidic chip 110 has three inlets, one inlet of the main channel 111 and two inlets of the auxiliary channel 112, and eight circular mixed herringbone structures 113 are arranged in the middle of the main channel 111, the microfluidic The chip 110 can realize the mixing of three liquids in continuous flow, and the liquid flowing to the outlet has been fully mixed.

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Abstract

本发明涉及生物医疗的技术领域，公开了一种用于质谱仪的上样装置，上样装置设置于质谱仪的进样口侧，上样装置包括微流芯片、连接管路、喷针和样品注入结构；样品注入结构将待测样品和反应溶液注入微流芯片；连接管路的一端连接于微流芯片的出口端；喷针连接于连接管路的另一端，喷针接通电源，通过电喷雾的方式使样品溶液形成喷雾；喷针产生喷雾喷射至质谱仪的进样口中，一方面该装置能与商品化质谱仪适配，实现在线样品高效混合-质谱进样，且混合后的反应时间精确调控，适用于特定生化反应中对不同时间点下样品动态过程实时的质谱表征；另一方面特殊的接口形式，连接紧密不漏液，离子化效率和灵敏度高，死体积小，便于更换。

Description

用于质谱仪的上样装置 技术领域
本发明涉及生物医疗的技术领域，具体涉及一种用于质谱仪的上样装置。
 背景技术
质谱（MS）是一种针对天然和非天然蛋白质的结构和动力学性质的有效方法。由于其高的分析灵敏度和特异性，MS在分子和药理学分析以及临床实践中具有显著优势。然而，这项技术仍需在如下方面提升性能： (1) 动态（时间）分辨能力。蛋白质在溶液环境下高级结构和相互作用具有高度的动态性，物种间可快速互变，还可形成大量中间体，而现有方法难以提供时间维度上的表征数据。(2) 操作性。部分方法步骤较多、对操作人员经验技术要求较高，在工业实践中进行大规模推广仍需化简。(3) 整合性。蛋白质表征所需的表征方面繁复，而一些现有技术只能各自为战，利用一类设备、一种方法或一套经验解决单一方面的表征问题，全面满足表征要求所需的经济及人力成本很大。尽管一些研究工作已经对方法整合进行了一些探索，但在集成化的目标上仍需做出更多硬件及方法上的努力。(4)样品制备步骤繁复、样品耗用量大。这些问题限制了质谱技术的进一步应用。例如传统质谱方法中，参与特定生化反应的样品需在特定时间节点进行反应淬灭后离线进样，难以实现较小时间尺度的时间分辨，且大量反应难以实现完全淬灭，后续处理及进样过程可引入预期外的进一步反应、副反应或逆反应，严重干扰测定结果。因此在样品在线反应、实时进样、动态调控以及自动化控制操作方面的技术提升可大幅拓展质谱技术的应用空间。
微流体设备在操纵和检测化学物质和生物分子方面具有各种优势。该平台可以实现低耗样量，可实现自动化、高通量操作以及温度控制，灵活的参数客户化以及可调控的反应时间，有了基于微流芯片的质谱上样装置，则可以在线完成样品前处理和生化反应片上操作，并在下游实时进样，从而能够精确检测特定反应时间条件下的产物特性，并且节省大量的样品浪费和人力成本，因此，微流芯片与质谱仪的结合是非常需要的。
 技术问题
其中，分析物离子化对于MS进样分析至关重要，微流芯片和质谱仪集成有诸多挑战，例如样品经前处理或反应后的总流速和溶液组成是否与质谱适配，如何对经前处理或反应的样品的高效离子化，以及将分析物从微流芯片转移到质谱仪的耦合界面。而玻璃或基于聚合物芯片的接口的制造以及可重复性并非易事，这也限制了在线的电喷雾质谱（ESI-MS）的普遍使用。
 技术解决方案
鉴于上述的不足之处，本发明的目的在于提供一种用于质谱仪的上样装置，该装置方案能在样品经前处理或反应后的总流速和溶液组成上与质谱适配，并且提供稳定且高离子化效率，并且芯片-质谱接口设计方便简易且可扩展。
为了实现上述目的，本发明提供的技术方案为：一种用于质谱仪的上样装置，所述上样装置设置于所述质谱仪的进样口侧，所述上样装置包括微流芯片、连接管路、喷针和样品注入结构；
所述样品注入结构将待测的样品溶液注入所述微流芯片；
所述连接管路的一端连接于所述微流芯片的出口端；
所述喷针连接于所述连接管路的另一端，所述喷针接通电源，通过电喷雾的方式使所述喷针内的样品溶液形成喷雾；
所述喷针产生喷雾喷射至所述质谱仪的进样口中。
进一步地，所述连接管路包括二通接头和套管，所述二通接头一端通过所述套管与所述微流芯片的出口端连接，所述二通接头的另一端通过所述套管与所述喷针连接。
进一步地，所述套管的材料为高分子聚合材料，所述套管与所述微流芯片的出口端过盈配合。
进一步地，所述微流芯片和所述喷针之间的夹角为锐角、直角或钝角，所述连接管路和所述喷针处于同一直线上。
进一步地，所述喷针的出口到所述质谱仪的进样口的距离为1-100mm。
进一步地，上文所述喷针包括但不限于不锈钢制的微孔套筒状结构。
进一步地，所述喷针可以简化为导电毛细管直接与所述微流芯片的出口商连接，无需二通接头、套管等零件。
进一步地，所述质谱仪包括电源，所述喷针与所述质谱仪的电源连接，由所述质谱仪提供电能。
进一步地，所述喷针的内径为1-200μm，外径为100-3000μm，长度为1-100mm  。
进一步地，所述微流芯片包括主流道和辅流道，所述主流道和辅流道用于加入不同的溶液；
所述辅流道连通于所述主流道，所述辅流道设置有多个。
进一步地，所述主流道中设置有导流结构，所述导流结构用于将所述主流道和所述辅流道中加入的不同溶液混合。
进一步地，所述导流结构包括鱼骨结构，所述鱼骨结构包括第一鱼骨结构和第二鱼骨结构，所述第一鱼骨结构和第二鱼骨结构为轴对称结构，所述第一鱼骨结构和第二鱼骨结构在所述主流道中间隔设置。
 有益效果
本发明通过用一个连接管路将喷针连接到微流芯片的出口端，再通过电喷雾技术将样品溶液喷射到质谱仪的进样口中，如此设置一方面可提高质谱仪的电喷雾效率和灵敏度，并改善对结构多样的分析物的鉴定；另一方面在结构上连接紧密，不漏液，在大流速通过的情况下，也可以保证不漏液，死体积很小，而且更换方便，简化了实验中需要更换微流芯片或喷针的操作，减少污染或喷针被堵时无法继续实验等的情况。
 附图说明
所包括的附图用来提供对本发明实施例的进一步的理解，其构成了说明书的一部分，用于例示本发明的实施方式，并与文字描述一起来阐释本发明的原理。显而易见地，下面描述中的附图仅仅是本发明的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动性的前提下，还可以根据这些附图获得其他的附图。在附图中：
图1是本发明的一实施例的一种上样装置和质谱仪的结构示意图；
图2是本发明的一实施例的连接管路和喷针的结构示意图；
图3是图1中局部A的剖视图；
图4是本发明的一实施例的微流芯片的结构示意图；
图5是本发明的一实施例的主流道和辅流道汇合处的结构和溶液混合的示意图；
图6是本发明的另一实施例的微流芯片的结构示意图。
其中，100、上样装置；110、微流芯片；111、主流道；112、辅流道；113、鱼骨结构；113a、第一鱼骨结构；113b、第二鱼骨结构；120、连接管路；121、二通接头；122、套管；130、喷针；140、样品注入结构；141、上样管路；142、溶液注射器；200、质谱仪；210、进样口；220、电源。
 本发明的实施方式
需要理解的是，这里所使用的术语、公开的具体结构和功能细节，仅仅是为了描述具体实施例，是代表性的，但是本发明可以通过许多替换形式来具体实现，不应被解释成仅受限于这里所阐述的实施例。
在本发明的描述中，术语“第一”、“第二”仅用于描述目的，而不能理解为指示相对重要性，或者隐含指明所指示的技术特征的数量。由此，除非另有说明，限定有“第一”、“第二”的特征可以明示或者隐含地包括一个或者更多个该特征；“多个”的含义是两个或两个以上。术语“包括”及其任何变形，意为不排他的包含，可能存在或添加一个或更多其他特征、整数、步骤、操作、单元、组件和/或其组合。
另外，“中心”、“横向”、“上”、“下”、“左”、“右”、“竖直”、“水平”、“顶”、“底”、“内”、“外”等指示的方位或位置关系的术语，是基于附图所示的方位或相对位置关系描述的，仅是为了便于描述本发明的简化描述，而不是指示所指的装置或元件必须具有特定的方位、以特定的方位构造和操作，因此不能理解为对本发明的限制。
此外，除非另有明确的规定和限定，术语“安装”、“相连”、“连接”应做广义理解，例如可以是固定连接，也可以是可拆卸连接，或一体地连接；可以是机械连接，也可以是电连接；可以是直接相连，也可以通过中间媒介间接相连，或是两个元件内部的连通。对于本领域的普通技术人员而言，可以根据具体情况理解上述术语在本发明中的具体含义。
下面参考附图和可选的实施例对本发明作进一步说明。
如图1所示，本发明实施例公开了一种用于质谱仪200的上样装置100，所述上样装置100设置于所述质谱仪200的进样口210侧，所述上样装置100包括微流芯片110、连接管路120、喷针130和样品注入结构140；所述样品注入结构140将待测的样品溶液注入所述微流芯片110；所述连接管路120的一端连接于所述微流芯片110的出口端；所述喷针130连接于所述连接管路120的另一端，所述喷针130接通电源220，通过电喷雾的方式使使所述喷针130内的样品溶液形成喷雾；所述喷针130产生喷雾喷射至所述质谱仪200的进样口210中。
具体的，通过电喷雾的方式使使所述喷针130内的样品溶液形成喷雾的原理为：细小的雾滴从金属喷针130喷射出来时，就从金属喷针130口的高强电场中获得了大量的电荷，由于受电荷之间库仑力的作用，这些电荷均匀地分布在液滴的表面。当液滴被干燥去溶剂时，液滴体积逐渐减小，于是单位表面积上的电荷急剧增加，使得液滴不稳定而进行分裂，产生更细小的液滴喷雾。
本实施例通过用一个连接管路120将喷针130连接到微流芯片110的出口端，再通过电喷雾技术将样品溶液喷射到质谱仪200的进样口210中，如此设置一方面可提高质谱仪200的电喷雾效率和灵敏度，并改善对结构多样的分析物的鉴定；另一方面在结构上连接紧密，不漏液，在大流速通过的情况下，也可以保证不漏液，死体积很小，而且更换方便，简化了实验中需要更换微流芯片110或喷针130的操作，减少污染或喷针130被堵时无法继续实验等的情况。
其中，所述质谱仪200包括电源220，所述喷针130与所述质谱仪200的电源220连接，由所述质谱仪200提供电。通过质谱仪200来给上样装置100供电来进行电喷雾，不需要再额外增加电源220，减小了上样装置100的体积，增加了上样装置100的便携性和使用的方便性。
进一步地，所述喷针130的出口到所述质谱仪200的进样口210的距离为1-100mm。保证样品溶液通过电喷雾形成的微小液滴能够均匀的飞到所述质谱仪200的进样口210中，防止距离过短喷入到质谱仪200进样口210中的微小液滴不均匀，或者距离过长导致有些形成的微小液滴不能飞到质谱仪200的进样口210中，掉在外面造成浪费或形成污染。
如图2和3所示，在一实施例中，所述连接管路120包括二通接头121和套管122，所述二通接头121一端通过所述套管122与所述微流芯片110的出口端连接，所述二通接头121的另一端通过所述套管122与所述喷针130连接。所述套管122为高分子聚合材料，所述套管122通过摩擦力，直接插入所述微流芯片110的出口端，与所述微流芯片110出口端过盈配合，连接紧密不漏液，即使在大流速通过下，也可以保证不漏液；在套管122堵住或需要清洗的时候，直接拔出进行更换或清洗即可，拆装方便，更换快速简单，减少操作时无法继续实验等的情况。
具体的，所述套管122的材料可以为PEEK聚醚醚酮、PEK聚醚酮、PEKK聚醚酮酮、PEEKK聚醚醚酮酮、PEKEKK聚醚酮醚酮酮、PFA 全氟烷氧基树脂、FEP氟化乙烯丙烯、ETFE乙烯-四氟乙烯或PTFE聚四氟乙烯等多种适合质谱实验分析的高分子聚合材料。
其中，所述喷针130为金属喷针130，所述喷针130的内径为1-200μm，外径为100-3000μm，长度为1-100mm；所述套管122的内径为150~200μm。具体地，所述喷针130的内径为30μm，外径为150μm，长度为40μm；所述套管122的内径为180μm。
进一步地，所述喷针130、套管122和二通接头121连接之后处于同一直线上，并且与所述微流芯片110的总体流动方向的夹角为锐角、直角或钝角。目前的连接方式都是使得加入的样品溶液流动的总体方向是一条直线，这样就需要比较精确的设计，密封也需要紧密的连接设计，在这种情况下，更换就很麻烦，还需要精准的调校，需要较长的时间，就会耽误样品溶液的检测或实验的进行。
如图4和5所示，在一实施例中，所述微流芯片110包括主流道111和辅流道112，所述样品注入结构140包括上样管路141和溶液注射器142，所述上样管路141分别插入所述主流道111和辅流道112的入口处，所述上样管路141的另一端连接溶液注射器142，通过溶液注射器142加压将样品溶液和添加溶液分别注入到所述主流道111和辅流道112中；所述辅流道112连通于所述主流道111，所述辅流道112设置有多个，每个辅流道112之间间隔11500μm。该微流芯片110可以实现多路液体的在线混合，在主流道111中加入主要的样品溶液，通过在辅流道112中加入所需的添加液体，在后续流道中完成混合，即在微流芯片110中即可完成溶液的混合和反应，连续流动操作，进行质谱上样分析，不需要先对溶液进行混合后再用来检测，要更改混合比例还需停止检测进行操作。并且通过溶液注射器142调节注入压力来调节溶液的流速，可以实时改变不同的混合比例，实现不同的反应混合比例、反应时间和混合效果的操作。在氢氘交换质谱实验中，通过流速控制，可以实时在线调节蛋白质氢氘交换反应时间，实现了在线动态分析蛋白质结构的操作。如图5所示，展示了主流道111（CH1）的溶液和辅流道112（CH2）的溶液的混合平衡效果，该微流芯片110可以实现从99:1到50:50的不同比例下的混合功能。
其中，所述主流道111中设置有导流结构，所述导流结构用于将所述主流道111和所述辅流道112中加入的不同溶液混合。具体的，所述导流结构为鱼骨结构113（Herringbone结构），按照溶液流动的方向，所述鱼骨结构113设置于所述主流道111与第一个所述辅流道112汇合之后的位置。在溶液流入鱼骨结构113处的时候会形成涡流或者混沌流，可以快速混合均匀，提高均匀混合的效率，缩短混合所需的流道的长度，从而可以使微流芯片110的大小缩减的更为小巧。当然，所述导流结构并不限制于鱼骨结构113，其他用于混合的几何结构也可以。
进一步地，所述鱼骨结构113包括第一鱼骨结构113a和第二鱼骨结构113b，所述第一鱼骨结构113a和第二鱼骨结构113b为轴对称结构，所述第一鱼骨结构113a和第二鱼骨结构113b在所述主流道111中间隔设置。顺着第一鱼骨结构113a和第二鱼骨结构113b的形状进行循环混合，使溶液的混合的更加均匀和更加快速。
具体的，所述微流芯片110包括一个主流道111和两个辅流道112，所述主流道111为一个主线流道，两个所述辅流道112分别接通到所述主流道111中，两个所述辅流道112之间的间隔为4000μm~11500μm，视循环数而定。一个第一鱼骨结构113a加一个第二鱼骨结构113b为一个循环，在所述主流道111中设置八个循环，即八个第一鱼骨结构113a和八个第二鱼骨结构113b间隔设置，即两个所述辅流道112之间的间隔设置为11500μm。当然，所述辅流道112的设置方式并不限于间隔设置，所述辅流道112之间还可以为分层的排布方式，或者是呈夹角设置。
在本实施例中，实现了质谱芯片研究中，在线进行蛋白质和反应溶液的边混合边上样的操作，并且混合快速有效且稳定，通过流速调节，还可以控制蛋白质反应时间，实时动态的进行质谱分析解析蛋白质结构。该微流芯片110可以实现多路液体的在线混合，在主流道111中加入主要的样品溶液，通过在辅流道112中加入所需的添加液体，通过循环的第一鱼骨结构113a加一个第二鱼骨结构113b快速均匀地将溶液进行混合和样品在线反应，连续流动操作，进行质谱上样分析。并且通过溶液注射器142调节注入压力来调节溶液的流速，可以实时改变不同的混合比例，实现不同的反应混合比例、反应时间和混合效果的操作。而且，液体在线混合比例范围和流速范围较大，可满足多种蛋白质的混合反应操作。
如图6所示，在另一实施例中，一个主流道111和两个辅流道112组成一组流道，所述微流芯片110包括独立的两组流道，当然，也可以设置多组流道，可以分多次使用并不需要清洗，也可以同时对不同的样品溶液进行检测。在微流芯片110制造时直接设计多组流道是比较容易的，对制造难度的影响也不大，这样可以节省原材料和时间。微流芯片材质可为玻璃石英、硅片、纸基材料和高分子聚合物（PDMS、PMMA和PC等）等。上述微流芯片110的制造方法具体为：使用制图软件进行设计具有鱼骨结构113的微流芯片110，并制作菲林，之后通过光刻技术制作微流芯片模板。示例性的微流芯片110用聚二甲基硅氧烷 (Polydimethylsiloxane，PDMS) A液和B液按照10:1混合均匀后，倒入前面制作出的光刻模板中，经过抽真空脱泡后，水平放置于80度烘箱，静置45分钟，脱模，得到对应PDMS芯片，用0.75 mm的PDMS打孔器分别在入口和出口初打通孔。用Plasma等离子清洗机将PDMS芯片键合在玻璃载玻片上，放入80度烘箱，放置2个小时或一晚上。
具体的，该微流芯片110有三个入口，一个主流道111的入口和两个辅流道112的入口，并且在主流道111的中间设置有8个循环的混合鱼骨结构113，该微流芯片110可以实现三路液体在连续流动中的混合，流到出口的液体已经得到充分的混合。
需要说明的是，本方案中涉及到的各步骤的限定，在不影响具体方案实施的前提下，并不认定为对步骤先后顺序做出限定，写在前面的步骤可以是在先执行的，也可以是在后执行的，甚至也可以是同时执行的，只要能实施本方案，都应当视为属于本发明的保护范围。
以上内容是结合具体的可选的实施方式对本发明所作的进一步详细说明，不能认定本发明的具体实施只局限于这些说明。对于本发明所属技术领域的普通技术人员来说，在不脱离本发明构思的前提下，还可以做出若干简单推演或替换，都应当视为属于本发明的保护范围。

Claims (10)

  1. 一种用于质谱仪的上样装置，其特征在于，所述上样装置设置于所述质谱仪的进样口侧，所述上样装置包括微流芯片、连接管路、喷针和样品注入结构；
    所述样品注入结构将待测的样品溶液注入所述微流芯片；
    所述连接管路的一端连接于所述微流芯片的出口端；
    所述喷针连接于所述连接管路的另一端，所述喷针接通电源，通过电喷雾的方式使所述喷针内的样品溶液形成喷雾；
    所述喷针产生喷雾喷射至所述质谱仪的进样口中。
  2. 根据权利要求1所述的用于质谱仪的上样装置，其特征在于，所述连接管路包括二通接头和套管，所述二通接头一端通过所述套管与所述微流芯片的出口端连接，所述二通接头的另一端通过所述套管与所述喷针连接。
  3. 根据权利要求2所述的用于质谱仪的上样装置，其特征在于，所述套管的材料为高分子聚合材料，所述套管与所述微流芯片的出口端过盈配合。
  4. 根据权利要求1所述的用于质谱仪的上样装置，其特征在于，所述微流芯片和所述喷针之间的夹角为锐角、直角或钝角，所述连接管路和所述喷针处于同一直线上。
  5. 根据权利要求1所述的用于质谱仪的上样装置，其特征在于，所述喷针的出口到所述质谱仪的进样口的距离为1-100mm。
  6. 根据权利要求1所述的用于质谱仪的上样装置，其特征在于，所述质谱仪包括电源，所述喷针与所述质谱仪的电源连接，由所述质谱仪提供喷雾电压。
  7. 根据权利要求1-6任意一项所述的用于质谱仪的上样装置，其特征在于，所述喷针的内径为1-200μm，外径为100-3000μm，长度为1-100mm。
  8. 根据权利要求1-6任意一项所述的用于质谱仪的上样装置，其特征在于，所述微流芯片包括主流道和辅流道，所述主流道和辅流道用于加入不同的溶液；
    所述辅流道连通于所述主流道，所述辅流道设置有多个。
  9. 根据权利要求8所述的用于质谱仪的上样装置，其特征在于，所述主流道中设置有导流结构，所述导流结构用于将所述主流道和所述辅流道中加入的不同溶液流动混合。
  10. 根据权利要求9所述的用于质谱仪的上样装置，其特征在于，所述导流结构包括鱼骨结构，所述鱼骨结构包括第一鱼骨结构和第二鱼骨结构，所述第一鱼骨结构和第二鱼骨结构为轴对称结构，所述第一鱼骨结构和第二鱼骨结构在所述主流道中间隔设置。
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