WO2014194537A1 - 一种可视化检测汗液中氯离子含量的便携式纸芯片 - Google Patents

一种可视化检测汗液中氯离子含量的便携式纸芯片 Download PDF

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WO2014194537A1
WO2014194537A1 PCT/CN2013/077471 CN2013077471W WO2014194537A1 WO 2014194537 A1 WO2014194537 A1 WO 2014194537A1 CN 2013077471 W CN2013077471 W CN 2013077471W WO 2014194537 A1 WO2014194537 A1 WO 2014194537A1
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paper
sweat
portable
color
volume
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PCT/CN2013/077471
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English (en)
French (fr)
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郑直
慕轩
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中国医学科学院基础医学研究所
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Priority to EP13886328.7A priority Critical patent/EP3006937B1/en
Publication of WO2014194537A1 publication Critical patent/WO2014194537A1/zh

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N31/00—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods
    • G01N31/22—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators
    • G01N31/221—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators for investigating pH value
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
    • A61B5/00—Measuring for diagnostic purposes; Identification of persons
    • A61B5/42—Detecting, measuring or recording for evaluating the gastrointestinal, the endocrine or the exocrine systems
    • A61B5/4261—Evaluating exocrine secretion production
    • A61B5/4266—Evaluating exocrine secretion production sweat secretion
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
    • G01N33/525—Multi-layer analytical elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
    • G01N33/525—Multi-layer analytical elements
    • G01N33/526—Multi-layer analytical elements the element being adapted for a specific analyte
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/84—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving inorganic compounds or pH
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/483—Physical analysis of biological material
    • G01N33/487—Physical analysis of biological material of liquid biological material

Definitions

  • the present invention relates to a paper-based, in situ, visually small device for detecting chloride ions in sweat, and its use for physiological monitoring and preliminary screening for cystic fibrosis, a hereditary disease. Background technique
  • Sweat is a major form of body fluids, including metal salts, sugars, organic acids and many other compounds. Its main features are: 1) direct excretion, not invasive collection, with very low risk of infection; 2) exercise, heat and compounds can stimulate the discharge of sweat, easy to obtain; 3) its chemical composition and specific Physiological and pathological conditions are relevant. If the body is dehydrated, the concentration of chloride ions in the sweat will increase significantly.
  • the traditional method for detecting chloride ions in sweat is: 1) using a pilocarpine iontophoresis to subcutaneously stimulate the sweat to flow out; 2) collecting the sweat by using a dry gauze; 3) weighing the gauze after the sweat is absorbed, and calculating the volume of sweat flowing out 4)
  • the atomic luminometer is used to detect the ion concentration in the eluent of the gauze.
  • gauze it is also possible to use a liquid storage bag and a cotton cloth as described in C.
  • the present invention provides a portable paper chip characterized in that it has paper having a color or fluorescence change depending on pH and an anion exchange paper disposed on the lower surface of the paper.
  • the portable paper chip according to the present invention further has a viscous sticker disposed on the upper surface of the paper which changes color or fluorescence according to pH, and a paper characterization of the liquid inlet volume disposed on the lower surface of the adhesive sticker.
  • the paper characterizing the incoming liquid volume has a liquid inlet end.
  • a portable paper chip further comprising a substrate disposed on a lower surface of the portable paper chip, the substrate and the adhesive sticker, the anion exchange paper, the paper that changes color or fluorescence according to pH
  • the position of the paper characterizing the liquid inlet volume is fixed therebetween, and has pores on the substrate corresponding to the position of the anion exchange paper and the liquid inlet end of the paper characterizing the liquid inlet volume.
  • the paper which changes color or fluorescence depending on pH is a pH test paper or a paper impregnated with a pH-responsive fluorescent dye
  • the substrate is a water-impermeable material, preferably a water-impermeable tape.
  • the anion exchange paper is paper modified with an anion exchange group, and the anion exchange group is preferably a diethylaminoethyl group or a quaternary ammonium group.
  • the paper tape characterizing the liquid inlet volume has a fluorescent or color marking and an indicator scale.
  • the adhesive sticker is transparent, translucent or has a visible window.
  • the invention provides the use of a portable paper chip according to the invention for detecting chloride ion content in sweat.
  • the invention provides an in situ, rapid detection of chloride ion content in sweat Method, which includes the following steps:
  • the portable paper chip according to the present invention is covered on the surface of the skin, and the sweat is sucked through the anion exchange paper;
  • the chloride ion content in the sweat is determined from the pre-rendered standard curve by the color or fluorescence change of the paper which changes color or fluorescence according to pH.
  • the invention provides a method for in situ, rapid detection of chloride ion content in sweat comprising the following steps:
  • the portable paper chip according to the present invention is applied to the surface of the skin, and the sweat is sucked through the anion exchange paper and the liquid inlet end of the paper characterization of the liquid volume;
  • the volume of the absorbed sweat is determined by the paper characterizing the volume of the influent, and when the volume of the inhaled sweat reaches a certain value, the portable paper chip is removed;
  • the chloride ion content in the sweat is determined based on a pre-rendered standard curve by color change of the paper which changes color or fluorescence according to pH.
  • the volume of sweat absorbed is preferably from 0.1 to 10 ⁇ L, more preferably 2 ⁇ L.
  • the paper chip can integrate functions such as sweat collection and detection, eliminating the transfer step of sweat, simplifying the operation steps and improving the accuracy of detection;
  • the paper chip can detect the chloride ions in the sweat in situ, real-time and visually, so as to monitor the physiological conditions;
  • the paper chip is mainly composed of paper, which is extremely portable and low in cost, and is very suitable for large-scale screening and daily use;
  • Paper chips use biosafety adhesive stickers that can be used on the skin.
  • 1 is a schematic exploded view of a portable paper chip for in-situ detection of chloride ions according to the present invention; wherein 1 is an impermeable material, 2 is an anion exchange paper, 3 is a paper that produces a color or fluorescence change according to pH, and 4 is a representation Into the volume of the paper, 5 is a paper with adhesive, 6 is a hole, 7 is a scale, 10 is the liquid inlet end of the paper characterizing the volume of the liquid;
  • 2 is a schematic cross-sectional view of a portable paper chip for in-situ detection of chloride ion content according to the present invention; wherein 1 is a water-impermeable material, 2 is an anion exchange paper, and 3 is a paper which produces a color or fluorescence change according to pH, 4 To characterize the paper volume, 5 is a sticky paper, 6 is a hole, and 7 is a scale;
  • Figure 3 is a schematic diagram of the mechanism for detecting chloride ions, wherein 2 is anion exchange paper, 3 is paper which produces color or fluorescence change according to pH, 8 is sweat, 9 is skin of healthy person or cystic fibrosis patient; 11 arrow indicates liquid Direction of flow;
  • Figure 4 shows the color change corresponding to different chloride ion concentrations.
  • Five circular holes correspond to five detection areas, the lower number is the concentration of chloride ions, and the color channel is Cyan in the CMYK color space.
  • Figure 5 shows the standard curve for 0 to 100 mmol chloride detection and the results for healthy people (triangles) and cystic fibrosis patients (diamonds).
  • the present invention relates to a visualized chloride ion detecting paper chip device fabricated by anion exchange paper and paper assembled by color or fluorescence depending on pH.
  • the paper is preferably a layered structural film composed of three-dimensional cellulose fibers, and is hydrophilic.
  • the anion exchange paper is a paper modified with an anion exchange group such as diethylaminoethyl and quaternary ammonium groups, capable of adsorbing anions in a liquid and exchanging hydroxide ions. Which produces color changes depending on pH
  • the paper is a pH test strip (such as a wide pH test strip and a precision pH test strip) that reflects the change in pH by color change.
  • the paper chip device of the present invention further has a paper characterizing the volume of the liquid having a scale and a color mark of fluorescein or a dye thereof, and when the liquid is sucked at the liquid inlet end, the mark indicates the scale of the liquid front end. , thereby indicating the volume of liquid absorbed by the paper. More preferably, the paper chip device of the present invention has a transparent and viscous sticker such as 3M tegaderm.
  • the sweat on the skin first passes through an anion exchange paper where the anions are exchanged for hydroxide ions.
  • the volume of sweat is estimated from paper that can indicate the volume of the influent.
  • the anion of the sweat is mainly chloride ions, which are generally several thousand times higher than the sulfate anions in the second position.
  • the pH of the sweat increases accordingly, and the magnitude of the change is related to the displaced hydroxide.
  • the anion exchange paper the sweat will come into contact with the pH test paper, and the color change on the pH test paper can indicate the change in pH.
  • pH-responsive fluorescent materials can also be used to indicate changes in pH. Color or fluorescence changes are captured by a CCD or smart phone and image processed to separate the Red channel in the RGB color space or the Cyan channel in the CMYK color space. The gray scale in a single channel color indicates the concentration of chloride ions in the sweat.
  • Step 1 Make the corresponding paper structure according to the design size, see Figure 1; 6 is a round hole with a diameter of 1-3 mm; 2 is an anion exchange paper, Whatman DE81 or Pall Mustang, its diameter is slightly larger than 6, 2 -4 mm, 3 is a pH test paper or a paper modified with pH-sensitive quantum dots (fluorescent dye), the diameter of which is slightly larger than 2, 3-5 mm; the diameter of the round end of 4 is slightly larger than 6 and is 2-4 Mm, 4 has a length of 2 cm; 1 has a length of 3-5 cm and a width of 1-3 cm; 7 is made by an inkjet printer; 5 is a 3M Tegaderm adhesive paper, which is slightly larger than 1 and has a length of 4 6 cm, width 2-4 cm;
  • Step 2 Assemble the 1-5 according to the design to obtain a paper chip. 5 has a viscosity, through the tight combination with 1 to fix the position of other components; the cross section of the paper chip after assembly is shown in Figure 2;
  • the third step preparing a gradient concentration solution of sodium chloride, 10 mM-80 mM;
  • Step 4 Add 1 ⁇ l of sodium chloride solution to the 6 in 1, let the liquid flow into 2, 3; exchange the chloride ions to hydroxide ions, raise the pH of the liquid, and change the color of the pH test paper. , see Figure 3;
  • Step 5 Use CCD or smart phone to capture the color change of pH test paper, see Figure 4;
  • the shooting background is generally pure blue, and the shooting light is evenly distributed;
  • Step 6 Separate the color information of the single channel, such as the R channel in RGB and the C channel in CMYK, with the chloride channel concentration by software (such as NIH ImageJ or Adobe Photoshop); establish a standard curve of chloride ion concentration.
  • software such as NIH ImageJ or Adobe Photoshop
  • Step 1 Make the corresponding paper structure according to the design size, see Figure 1; Make the corresponding paper structure according to the design size, and assemble the corresponding paper structure; 6 diameter is 1-3 mm, 2 diameter is slightly larger than 6 2-4 mm, the diameter of 3 is slightly larger than 2, 3-5 mm; the diameter of the round end of 4 is slightly larger than 6 and is 2-4 mm, the length of 4 is 2 cm; the length of 1 is 3-5 Cm, width 1-3 cm; 5 to 1 slightly larger, 4-6 cm long, 2-4 cm wide;
  • Step 2 Assemble the 1-5 according to the design to get the paper chip. 5 has adhesiveness, and fixes the position of other parts by tightly bonding with 1; the cross section of the paper chip after assembly is shown in Fig. 2;
  • the third step preparing a gradient concentration solution of sodium chloride, O mM-lOO mM;
  • Step 4 Detect a sodium chloride solution to establish a standard curve; place the paper chip on 9 and wait for the sweat to enter the paper chip;
  • Step 5 When the scale is indicated to 2 ⁇ l, remove the paper chip, or directly use the CCD or smart phone to collect the image information of the pH test paper;
  • Step 6 Use software (such as smartphone app, NIH ImageJ or Adobe)

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Abstract

一种可视化检测汗液中氯离子含量的便携式纸芯片，具有根据pH产生颜色或荧光变化的纸（3）以及在该纸下表面设置的阴离子交换纸（2）的双层结构。使用该纸芯片可对汗液中氯离子含量进行可视化检测，进而监控生理情况和诊断囊性纤维化疾病。

Description

一种可视化检测汗液中氯离子含量的便携式纸芯片 技术领域
本发明涉及一种基于纸的原位、 可视化检测汗液中氯离子的小型 设备， 以及其用于生理情况监控和初步筛查囊性纤维化 （一种遗传性 疾病） 的用途。 背景技术
可视化、 便携式的体液检测设备是个人化医疗未来发展的重要方 向之一。 便携式的设备可以允许在家庭和公共场所进行临床检测， 而 可视化的检测结果可以快速反映体液中的生理和病理信息， 极大地提 高医疗诊断的效率。 汗液是一种主要的体液形式， 包括金属盐、 糖、 有机酸等多种化合物。它主要的特点是 1 )直接排出体外， 不是侵入式 的收集方式， 具有非常低的感染风险； 2)运动、 热源和化合物都可以 刺激汗液的排出， 获取方便； 3 ) 其化学成分与特定的生理、 病理情况 具有相关性。 如身体在脱水的情况下， 汗液中氯离子的浓度就会显著 提高。 通过监控汗液中的氯离子浓度就可以获知身体的脱水情况， 从 而促进有效补水， 防止脱水导致的慢性和急性疾病。 另外， 患有囊性 纤维化 （Cystic Fibrosis ) 病人的一种跨膜转运蛋白会产生突变， 无法 从汗液中回收氯离子， 因此也会造成汗液中氯离子浓度的提高。 通过 检测汗液中的氯离子浓度已经成为诊断囊性纤维化的主要手段和金标 准。
尽管检测汗液中的氯离子具有重要的临床意义和实用价值， 但是 相关检测技术和设备的发展却十分不足， 尤其是缺乏便携式、 可原位 检测的能力。汗液中氯离子的传统检测方法的流程是 1 )用毛果芸香碱 电离子导入皮下刺激汗液流出； 2 ) 使用称重干燥纱布收集流出汗液； 3 ) 称重吸收汗液后纱布的重量， 计算流出汗液的体积； 4 ) 再用原子 发光光度计检测纱布的洗脱液中离子浓度。 除了纱布， 还可以使用如 C. Palacios, K等人在 Sweat mineral loss from whole body, patch and arm bag in white and black girls. Nutrition Research 23, 401 (Mar, 2003)中描 述的储液袋和棉布贴来收集汗液。 目前最常用的商品化仪器是由美国 Wescor公司研发的 Macroduct System ( Wescor Inc., http://www.wescor.com/biomedical/cysticfibrosis/ macroduct.html禾口 M. R. Ely等人， Evaluation of the Megaduct sweat collector for mineral analysis. Physiological Measurement 33, 385 (Mar, 2012))。该系统也获得了美国囊性纤维化协会的认可，可以用于囊性纤 维化疾病的诊断。 但是这些方法最大的问题在于汗液收集与汗液检测 步骤相对独立， 因此收集和检测之间仍然存在汗液转移步骤， 不仅不 能得到原位实时的检测结果，还容易产生误差。 M. Constantinescu, B. C. Hilman, The sweat test for quantitation of electrolytes - A challenge in precision. Laboratory Medicine 27, 472 (Jul, 1996)的研究报道称， 滤纸和 棉布上收集的汗液每分钟会因为蒸发损失 2%的体积。此外， Macroduct System系统结构复杂， 检测流程繁琐， 价格也较昂贵， 限制了其进一 步的应用。
除了以上收集和检测独立进行的技术， 汗液中氯离子的原位检测 方法也有报道。 一些早期的研究利用铬酸银与氯离子生成氯化银沉淀 的反应来检测汗液中的氯离子 （参见， J. B. J. S. S. J. Knights Em, Simplified screening test for cystic fibrosis of the pancreas. JAMA: The Journal of the American Medical Association 169, 1279 (1959)； H. Shwachman, N. Gahm, Studies in Cystic Fibrosis of the Pancreas. N. Engl. J. Med. 255, 999 (1956)； L. Gluck, A PATCH TEST FOR CHLORIDE IN SWEAT AS A SIMPLE SCREENING METHOD FOR DETECTING CYSTIC FIBROSIS OF THE PANCREAS: Preliminary Report. Pediatrics 23, 731 (April 1, 1959, 1959))。 虽然这种检测可以通过把出汗的手掌放 在琼脂糖凝胶和滤纸上进行， 操作简便， 结果直观， 但是铬酸银具有 一定的毒性和致癌性。 美国囊性纤维化协会已经明确禁止在皮肤上使 用基于氯化银沉淀的检测反应 （参见， V. A. LeGrys, J. R. Yankaskas, L. M. Quittell, B. C. Marshall, P. J. McGayzel, Diagnostic sweat testing: The cystic fibrosis foundation guidelines. Journal of Pediatrics 151, 85 (Jul,
2007))。最近， 有研究组发展出可佩戴的装置来检测汗液中氯离子（参 见， B. Schazmann 等人, A wearable electrochemical sensor for the real-time measurement of sweat sodium concentration. Analytical Methods 2, 342 (Apr, 2010))。 该装置主要由电子万用表和氯离子选择性电极组 成， 不仅采样量较大 （即需要大量的汗液）， 使应用对象和场合都受到 限制， 而且， 其体积和重量都严重影响了便携式的效果。
公开于该发明背景技术部分的信息仅仅旨在加深对本发明的一般 背景技术的理解， 而不应当被视为承认或以任何形式暗示该信息构成 已为本领域技术人员所公知的现有技术。 发明内容
本发明提供了一种便携式纸芯片， 其特征在于， 其具有根据 pH产 生颜色或荧光变化的纸以及在该纸下表面设置的阴离子交换纸。
进一步地， 根据本发明的便携式纸芯片， 其进一步具有设置于所 述根据 pH产生颜色或荧光变化的纸上表面的粘性贴纸以及在该粘性 贴纸的下表面设置的表征进液体积的纸， 所述表征进液体积的纸具有 进液端。
根据本发明的便携式纸芯片， 其进一步具有在所述便携式纸芯片 的下表面设置的基底， 所述基底与所述粘性贴纸将所述阴离子交换纸、 所述根据 pH产生颜色或荧光变化的纸、表征进液体积的纸的位置固定 在其中间， 并且在所述基底上对应于阴离子交换纸和表征进液体积的 纸的进液端的位置具有孔。
根据本发明便携式纸芯片， 优选地， 所述根据 pH产生颜色或荧光 变化的纸是 pH试纸或浸润 pH响应荧光染料的纸， 所述基底是不透水 材料， 优选为不透水胶带。
根据本发明的便携式纸芯片， 优选地， 所述阴离子交换纸是修饰 有阴离子交换基团的纸， 阴离子交换基团优选为二乙氨基乙基或季铵 基。
根据本发明的便携式纸芯片， 优选地， 所述表征进液体积的纸带 有荧光或颜色标记和指示刻度。
根据本发明的便携式纸芯片， 优选地， 所述粘性贴纸是透明的， 半透明的或者具有可视窗口的。
另一方面， 本发明提供了根据本发明的便携式纸芯片在检测汗液 中氯离子含量的用途。
另一方面， 本发明提供了一种原位、 快速检测汗液中氯离子含量 的方法， 其包括以下步骤：
将根据本发明的便携式纸芯片覆盖于皮肤表面， 通过阴离子交换 纸吸取汗液；
通过根据 pH产生颜色或荧光变化的纸的颜色或荧光变化，根据预 先绘制的标准曲线， 确定汗液中氯离子含量。
在一方面， 本发明提供了一种原位、 快速检测汗液中氯离子含量 的方法， 其包括以下步骤：
将根据本发明的便携式纸芯片覆盖于皮肤表面， 通过阴离子交换 纸和表征进液体积的纸的进液端吸取汗液；
通过表征进液体积的纸确定吸取汗液的体积， 当吸取汗液的体积 达到确定的数值时， 取下便携式纸芯片；
通过根据 pH产生颜色或荧光变化的纸的颜色变化，根据预先绘制 的标准曲线， 确定汗液中氯离子含量。
根据本发明的方法， 优选地， 吸取汗液的体积优选为 0.1至 10微 升， 更优选为 2微升。
通过纳入本文的附图以及随后与附图一起用于说明本发明的某些 原理的具体实施方式， 本发明的方法和装置所具有的其它特征和优点 将变得清楚或更为具体地得以阐明。
本发明的有益效果主要有以下几点：
纸芯片能够集成汗液收集和检测等功能， 免除了汗液的转移步骤， 简化操作步骤， 提高检测的准确性；
纸芯片可以对汗液中的氯离子进行原位、 实时和可视化检测， 方 便对生理情况的监测；
纸芯片主要由纸组成， 具有极高的便携型和低廉的成本， 非常适 用于大规模筛查和日常使用；
纸芯片使用生物安全性的粘性贴纸， 可以在皮肤上使用。 附图说明
以下， 结合附图来详细说明本发明的实施方案， 应当了解， 以下 技术方案仅用于说明本发明， 并不能限制本发明的范围， 本领域技术 人员可在不偏离本发明主旨的基础上， 对其进行任何变型。 其中： 图 1 为本发明所述用于原位检测氯离子的便携式纸芯片的分解示 意图； 其中 1为不透材料， 2为阴离子交换纸， 3为根据 pH产生颜色 或荧光变化的纸， 4为表征进液体积的纸， 5为带有粘性的纸， 6为空 孔， 7为刻度， 10为表征进液体积的纸的进液端；
图 2为本发明的用于原位检测氯离子含量的便携式纸芯片的横截 面示意图； 其中， 1 为不透水材料， 2为阴离子交换纸， 3为根据 pH 产生颜色或荧光变化的纸， 4为表征进液体积的纸， 5为带有粘性的纸， 6为空孔， 7为刻度；
图 3为检测氯离子的机理示意图， 其中 2为阴离子交换纸， 3为根 据 pH产生颜色或荧光变化的纸， 8为汗液， 9为健康人或囊性纤维化 病人的皮肤； 11箭头表示液体流动的方向；
图 4为不同氯离子浓度对应的颜色变化， 5个圆孔对应 5个检测区 域， 下方的数字是氯离子的浓度， 颜色通道是 CMYK色彩空间中的 Cyan;
图 5为 0到 100 mmol氯离子检测的标准曲线， 以及对健康人（三 角） 和囊性纤维化病人 （菱形） 的检测结果。
应当了解， 所附附图并非按比例地显示了本发明的基本原理的图 示性的各种特征的略微简化的画法。 本文所公开的本发明的具体设计 特征包括例如具体尺寸、 方向、 位置和外形将部分地由具体所要应用 和使用的环境来确定。
在这些图形中， 贯穿附图的多幅图形， 附图标记引用本发明的同 样的或等同的部分。
以下具体实施方式仅用于说明本发明， 并没有将本发明限于该具 体实施方式的意图。 具体实施方式
本发明涉及通过阴离子交换纸和根据 pH产生颜色或荧光变化的 纸组装制作成的可视化氯离子检测纸芯片装置。 在本发明中， 纸优选 为由三维纤维素纤维构成的层状结构薄膜， 具有亲水性。 其中阴离子 交换纸是修饰阴离子交换基团 (如二乙氨基乙基和季铵基)的纸，能够吸 附液体中的阴离子， 并交换出氢氧根离子。其中根据 pH产生颜色变化 的纸是 pH试纸（如广泛 pH试纸和精密 pH试纸）， 其能够通过颜色变 化反映出 pH的变化。 根据 pH产生荧光变化的纸是由修饰有 pH敏感 的量子点和荧光染料的纸代替。 优选地， 本发明的纸芯片装置进一步 地具有表征进液体积的纸， 其具有刻度和荧光素或其染料的颜色标记， 用该纸的进液端吸液体时， 标记会指示液体前端的刻度， 从而指示纸 吸收液体的体积。 更优选地， 本发明的纸芯片装置具有透明且具有粘 性的贴纸， 如 3M tegaderm。
利用微流控纸芯片技术将这些功能型纸进行剪裁、 立体组装和集 成， 以构建完整的微流控纸芯片， 实现可视化检测氯离子的功能。 皮 肤上的汗液首先经过阴离子交换纸， 其中的阴离子被交换成氢氧根离 子。 汗液的体积由可以指示进液体积的纸来估算。 需要说明的是， 汗 液中的阴离子主要氯离子， 其含量一般比第二位的硫酸根阴离子高数 千倍。 阴离子被交换成氢氧根后， 汗液的 pH值会相应增加， 其改变幅 度与置换出的氢氧根有关系。经过阴离子交换纸， 汗液会与 pH试纸接 触， pH试纸上的颜色变化可以指示 pH值的变化。 类似的， pH响应的 荧光材料也可以用来指示 pH值的变化。 颜色或荧光变化由 CCD或智 能手机采集， 经过图像处理， 分离出 RGB色彩空间中的 Red通道或者 CMYK色彩空间中的 Cyan通道。采用单通道颜色的灰度指示出汗液中 的氯离子浓度。 实施例 1：
第一步： 按照设计尺寸制作相应的纸结构， 见图 1 ; 6为圆孔， 其 直径为 1-3 mm; 2为阴离子交换纸， Whatman DE81或 Pall Mustang, 其直径比 6略大， 2-4 mm, 3为 pH试纸或修饰有 pH敏感量子点 （荧 光染料） 的纸， 其的直径比 2略大， 3-5 mm; 4的圆形端直径比 6略 大， 为 2-4 mm, 4的长度为 2 cm; 1的长为 3-5 cm, 宽为 1-3 cm; 7 由喷墨打印机制作； 5为 3M Tegaderm粘性纸， 其尺寸比 1略大， 长为 4-6 cm, 宽为 2-4 cm;
第二步： 将 1-5按照设计组装起来， 得到纸芯片。 5具有粘性， 通 过与 1 的紧密结合， 固定其他部件的位置； 组装后纸芯片的横截面见 图 2; 第三步： 配制氯化钠的梯度浓度溶液， 10 mM-80 mM;
第四步：将 1微升的氯化钠溶液滴加在 1中的 6上，让液体流入 2、 3； 氯离子交换成氢氧根离子， 使液体的 pH值上升， 改变 pH试纸的 颜色， 见图 3;
第五步： 采用 CCD或智能手机捕捉 pH试纸的颜色变化， 见图 4; 拍摄背景一般为纯蓝色， 拍摄灯光均匀分布；
第六步： 用软件（如 NIH ImageJ或 Adobe Photoshop)分离出单通 道的颜色信息， 如 RGB中的 R通道和 CMYK中的 C通道， 与氯离子 浓度进行关联； 建立氯离子浓度的标准曲线。 实施例 2:
第一步： 按照设计尺寸制作相应的纸结构， 见图 1 ; 按照设计尺寸 制作相应的纸结构， 将相应的纸结构进行组装； 6的直径为 1-3 mm, 2 的直径比 6略大， 2-4 mm, 3的直径比 2略大， 3-5 mm; 4的圆形端直 径比 6略大， 为 2-4 mm, 4的长度为 2 cm; 1的长为 3-5 cm, 宽为 1-3 cm; 5比 1略大， 长为 4-6 cm, 宽为 2-4 cm;
第二步： 将 1-5按照设计组装起来， 得到纸芯片。 5具有粘性， 通 过与 1 的紧密结合， 固定其他部件的位置； 组装后纸芯片的横截面见 图 2;
第三步： 配制氯化钠的梯度浓度溶液， O mM-lOO mM;
第四步： 检测氯化钠溶液建立标准曲线； 将纸芯片放在 9上， 等 待汗液进入纸芯片；；
第五步： 待刻度指示到 2微升时， 取下纸芯片， 或直接采用 CCD 或智能手机采集 pH试纸的图像信息；
第六步： 用软件 （如智能手机的 app， NIH ImageJ 或 Adobe
Photoshop) 分离出单通道的颜色信息， 如 RGB色彩空间中的 Red通 道和 CMYK色彩空间中的 Cyan通道， 利用标准曲线， 得到汗液中氯 离子的浓度， 见图 5。

Claims

权利要求书:
1、 一种便携式纸芯片， 其特征在于， 其具有根据 pH产生颜色或 荧光变化的纸以及在该纸下表面设置的阴离子交换纸的双层结构。
2、 根据权利要求 1所述的便携式纸芯片， 其进一步具有设置于所 述根据 pH产生颜色或荧光变化的纸上表面的粘性贴纸以及在该粘性 贴纸的下表面设置的表征进液体积的纸， 所述表征进液体积的纸具有 进液端。
3、 根据权利要求 2所述的便携式纸芯片， 其进一步具有在所述便 携式纸芯片的下表面设置的基底， 所述基底与所述粘性贴纸将所述阴 离子交换纸、所述根据 pH产生颜色或荧光变化的纸、表征进液体积的 纸的位置固定在其中间， 并且在所述基底上对应于阴离子交换纸和表 征进液体积的纸的进液端的位置具有孔。
4、 根据权利要求 1至 3任一项所述的便携式纸芯片， 其中所述根 据 pH产生颜色或荧光变化的纸是 pH试纸或浸润 pH响应荧光染料的 纸， 所述基底是不透水材料， 优选为不透水胶带。
5、 根据权利要求 1至 4任一项所述的便携式纸芯片， 其中所述阴 离子交换纸是修饰阴离子交换基团的纸， 阴离子交换基团优选为二乙 氨基乙基或季铵基。
6、 根据权利要求 1至 5任一项所述的便携式纸芯片， 其中所述表 征进液体积的纸带有荧光或颜色标记和指示刻度。
7、 根据权利要求 1至 6任一项所述的便携式纸芯片， 其中所述粘 性贴纸是透明的， 半透明的或者具有可视窗口的。
8、 根据权利要求 1至 7任一项所述的便携式纸芯片在检测汗液中 氯离子含量的用途。
9、 一种原位、 快速检测汗液中氯离子含量的方法， 其包括以下步 骤：
将根据权利要求 1至 7任一项所述的便携式纸芯片覆盖于皮肤表 面， 通过阴离子交换纸吸取汗液；
通过根据 pH产生颜色或荧光变化的纸的颜色或荧光变化，根据预 先绘制的标准曲线， 确定汗液中氯离子含量。
10、 一种原位、 快速检测汗液中氯离子含量的方法， 其包括以下 步骤：
将根据权利要求 2至 7任一项所述的便携式纸芯片覆盖于皮肤表 面， 通过阴离子交换纸和表征进液体积的纸的进液端吸取汗液；
通过表征进液体积的纸确定吸取汗液的体积， 当吸取汗液的体积 达到确定的数值时， 取下便携式纸芯片；
通过根据 pH产生颜色或荧光变化的纸的颜色变化，根据预先绘制 的标准曲线， 确定汗液中氯离子含量。
11、 根据权利要求 10的方法， 其中吸取汗液的体积优选为 0.1至 10微升， 更优选为 2微升。
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CN103336007B (zh) 2016-04-13
EP3006937A4 (en) 2017-01-25
EP3006937A1 (en) 2016-04-13

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