CN100480695C - Electrochemical screen printing electrode sensing test piece and manufacturing method thereof - Google Patents

Abstract

The invention relates to an electrochemical screen printing electrode sensing test piece and a preparation method thereof. The test piece has the characteristics of quickly absorbing the sample, effectively controlling the volume of the detected sample and filling and positioning the sample. The test piece has a structure that two or three electrodes, namely a working electrode, an auxiliary electrode and a reference electrode (a three-pole type) are arranged on an insulating bottom plate; directly adsorbing a reaction layer on the surface of the electrode area; the reaction layer is composed of buffer solution, reaction catalyst, electron transfer substance, wetting agent and surfactant; a specimen diversion area is arranged above the electrode area, which is formed by combining an intermediate layer sheet with a U-shaped channel and an upper cover sheet thereon. The hollow groove has capillary and siphon phenomena, can quickly suck a very small amount of sample into the electrode reaction area, and then measures the reaction between the sample and the reaction catalytic substance and the electron transfer substance in the reaction area by an electrochemical constant potential method or a constant current method, thereby analyzing the target component in the quantitative sample. The upper cover plate is provided with a closed convex space at the front end of the electrode area adjacent to the lead, and has the functions of effectively controlling the volume of the sample and filling and positioning the sample.

Description

Translated fromChinese

一种电化学网版印刷电极感测试片及其制法A kind of electrochemical screen printing electrode sense test piece and its preparation method

技术领域technical field

本发明关于一种电化学或网版印刷电极感测试片及其制法。该试片利用毛细虹吸的原理，可以将极少量检体迅速吸入电极反应区，利用定电位法或定电流法进行检体成分的定量分析。The invention relates to an electrochemical or screen printing electrode sensing test piece and its preparation method. Using the principle of capillary siphon, the test piece can quickly suck a very small amount of sample into the electrode reaction area, and use the constant potential method or constant current method to carry out quantitative analysis of the sample components.

背景技术Background technique

近年来电极式检验试片由于其容易制造、成本低廉以及方便携带的价廉检测仪的普及，已被成功地应用于各种产品检测商业化生产；例如血糖、尿酸、胆固醇等各种生化检测器。以市场最大且应用最多的血糖检测仪为例，市面上技术领导厂商包括罗氏(Roche)、亚培(Abbott)、Bayer及Therasense等公司，都是以电化学式方式生产血糖检测试片。第一代电化学式血糖测试片的血量需求高(5-10μl以上)且时间亦长(30-60秒不等)，虽在血量及检测时间方面已胜过光学显色法(colorimetric)等传统方法，但在使用上仍并不理想。随着技术的进步，目前最新一代产品，血量只要0.3μl(Therasense公司Freestyle产品)或1μl(Lifescan公司oneTouch Ultra产品)，所需时间也已缩短至5-10秒，因此采血量少以及检测速度快，已成为此类产品与技术发展的方向，而且各种的电极结构的研发也都朝此方向发展。Diebold等人于1995年的美国专利5,437,999号，披露了一种利用对掌式电极设计并在两电极间作出一毛细导引区及向上开口的圆形导气口，检体可以精确小量的进入反应区作用。该发明可采用光罩式及网版印刷制程，但不论何种制法其组成都是由两片含单股电极基版对向夹制而成，不仅步骤繁琐，成本亦很高。Shieh于1998年的美国专利5,779,867号，亦披露了一种电极式血糖试片设计，该发明利用上下两片对应式电极，中间夹层为反应区并留有一圆孔做为检体滴入口，夹层区内含一血球分离薄膜以过滤红血球，本方法以上下两试片所夹薄膜来控制血量并期以分离薄膜去除血球的干扰，但因过滤膜的阻隔无法在血量与速度上达到最佳状态。亚培公司(美国专利6，129,823)利用单层或多层网层覆盖在工作电极表面，最上层再盖一层上盖，此上盖在工作电极正上方或邻近工作电极处开一孔洞，做为检体注入的位置。此设计亦达到只需要2.0-2.5μl小体积检体的目的。不过此发明是以网层作为减少血量以及分布检体的方法。Therasense公司于美国专利6,299,757及6,338,790两专利中披露了同样利用对掌式电极设计，在两电极间透过精细的构装，放入一片具有极佳吸水性的薄片，将检体吸至两电极间进行反应。利用严密的吸水薄膜体积控制，可将检体所需最小体积降至0.3μl，为目前全世界血量最少的设计，但该发明则在封装程序较复杂的情况下有成本极高的弱点。沈燕士先生等人亦于台湾专利208,095号中披露网版印刷电极的制法及其应用，其中强调先以网版印刷完成导电膜及电绝缘层，再以电镀方法完成金属层，并以绝缘胶于工作及参考电极处形成一圆形凹陷部，并于此凹陷部内吸附所谓的生物层，检测时直接将检体滴入此处，约需检体10μl。此方法需电镀等繁琐步骤且血量需求亦较高.另外五鼎公司于台湾专利124,332号中则披露于电极区上方作成一导流区，上方并覆盖一片含有表面活性剂的网膜，利用毛细与虹吸原理将检体吸至电极上反应，该法如同亚培的方法一般以覆盖的网膜做为导流的设计，除增加成本外亦受最小检体需求量的限制。In recent years, electrode-type test strips have been successfully used in commercial production of various product testing due to their easy manufacture, low cost, and the popularity of cheap detectors that are easy to carry; such as various biochemical tests such as blood sugar, uric acid, and cholesterol. device. Take the largest and most widely used blood glucose tester as an example. The technology leaders in the market, including Roche, Abbott, Bayer, and Therasense, all produce blood glucose test strips electrochemically. The first generation of electrochemical blood glucose test strips requires high blood volume (above 5-10 μl) and takes a long time (ranging from 30-60 seconds), although it has surpassed the colorimetric method in terms of blood volume and detection time And other traditional methods, but still not ideal in use. With the advancement of technology, the latest generation of products currently only need 0.3 μl of blood (Freestyle product of Therasense Company) or 1 μl (oneTouch Ultra product of Lifescan Company), and the required time has also been shortened to 5-10 seconds. Fast speed has become the development direction of such products and technologies, and the research and development of various electrode structures are also developing in this direction. Diebold et al.’s U.S. Patent No. 5,437,999 in 1995 disclosed a design that utilizes palm-type electrodes and creates a capillary guide area between the two electrodes and a circular air guide port that opens upward, so that the specimen can enter in a small amount precisely. The effect of the reaction zone. The invention can adopt photomask and screen printing process, but no matter what kind of production method, its composition is made by sandwiching two substrates with single-strand electrodes facing each other, which is not only cumbersome in steps, but also high in cost. Shieh's U.S. Patent No. 5,779,867 in 1998 also disclosed a design of an electrode-type blood glucose test strip. The invention uses two corresponding electrodes on the upper and lower sides. There is a blood cell separation membrane in the area to filter red blood cells. In this method, the blood volume is controlled by the membrane sandwiched between the upper and lower test pieces, and the separation membrane is used to remove the interference of blood cells. However, due to the barrier of the filter membrane, the blood volume and speed cannot be maximized. good condition. Abbott (U.S. Patent No. 6,129,823) covers the surface of the working electrode with a single or multi-layer mesh layer, and the uppermost layer is covered with a top cover. The top cover has a hole directly above the working electrode or adjacent to the working electrode. As the sample injection position. This design also achieves the purpose of only requiring a small volume of 2.0-2.5 μl. However, this invention uses the mesh layer as a method for reducing blood volume and distributing samples. Therasense company disclosed in US patents 6,299,757 and 6,338,790 that it also uses the palm-type electrode design, and puts a thin sheet with excellent water absorption between the two electrodes through a fine structure to suck the specimen to the two electrodes react in between. Using strict water-absorbing film volume control, the minimum volume required for the sample can be reduced to 0.3 μl, which is currently the design with the least blood volume in the world. However, this invention has the weakness of extremely high cost in the case of complicated packaging procedures. Mr. Shen Yanshi and others also disclosed the preparation method and application of screen printing electrodes in Taiwan Patent No. 208,095, which emphasizes that the conductive film and the electrical insulating layer are completed by screen printing first, and then the metal layer is completed by electroplating, and the insulating glue is used to complete the electrode. A circular depression is formed at the working and reference electrodes, and the so-called biological layer is adsorbed in this depression. The sample is directly dropped here during detection, and about 10 μl of the sample is required. This method requires cumbersome steps such as electroplating and requires high blood volume. In addition, Wuding Company disclosed in Taiwan Patent No. 124,332 that a diversion area is made above the electrode area, and the top is covered with an omentum containing a surfactant. The principle of capillary and siphon draws the sample to the electrode for reaction. This method, like Abbott's method, uses the covered omentum as a diversion design. In addition to increasing the cost, it is also limited by the minimum sample demand.

Winarta等人于2001年美国专利6,258,229号中披露一种抛弃式检测装置，宣称只需小于1μl的液态检体.其制法为在一长条片金/聚苯乙烯(gold/polyester)或氧化锡/金/聚苯乙烯(tin oxide/gold/polyester)表层的一端以二氧化碳雷射(CO₂ laser)，蚀刻此长条片以形成如同三电极式的三互不相传导的区块；在如同工作电极上面，沿着试片的前端粘贴上一含有U型开口的中间层，此时工作电极位于U型槽内，再于中间层上粘贴一含有一方形通气孔洞的上盖.因此在工作电极上端形成了一个由液体入口至通气孔中间的液体通道，此液体通道限制了所能承受的液体体积小于1μl。此设计一如Nankai等人早于1992年美国专利5,120,420中所披露的，只是两者导电层中电极制法不同。Nankai等人所披露的电极试片制法是以网版印刷的方法在一绝缘背版上刷制双电极形式的导电层，其液体通道组成的方式之一，是在工作电极处前后两端横贴上两片间隔层，于其上再贴上一片不含孔洞的上盖，如此形成了一横向通过工作电极的开口通道。以该方法检体进入通道的使用量则无法控制，而且会从排气孔流出造成污染。另一种组成方式，如上所述，Winarta等人的设计，即在工作电极上端贴上一含有U型开口的中间层，再于其上贴上一片中间含有一个通气孔洞的上盖，于是形成了一个由液体入口至通气口中间的液体通道。此种结构，当通气口过小，仍有检体由通气口流出的顾虑；当通气口适中，检体则会恰巧停留在通气口边缘，尤其在讲求试片缩小化的今天，以手取试片时，很容易因不小心碰触通气口导致检体外流而造成污染。Winarta et al disclosed a disposable detection device in US Patent No. 6,258,229 in 2001, claiming that only less than 1 μl of liquid specimen is required. One end of the surface layer of tin/gold/polystyrene (tin oxide/gold/polyester) is etched with carbon dioxide laser (CO₂ laser) to form three mutually non-conductive blocks like three electrodes; As on the working electrode, paste an intermediate layer containing a U-shaped opening along the front end of the test piece. At this time, the working electrode is located in the U-shaped groove, and then paste an upper cover containing a square ventilation hole on the intermediate layer. Therefore, in A liquid channel is formed on the upper end of the working electrode from the liquid inlet to the middle of the vent hole, and the liquid channel limits the volume of the liquid that can be tolerated to less than 1 μl. This design is the same as that disclosed by Nankai et al. in US Patent No. 5,120,420 in 1992, except that the electrodes in the conductive layer are made differently. The method for preparing the electrode test piece disclosed by Nankai et al. is to brush a conductive layer in the form of a double electrode on an insulating back plate by screen printing. One of the ways in which the liquid channel is formed is at the front and rear ends of the working electrode. Two spacer layers are pasted horizontally, and a top cover without holes is pasted on it, so that an open channel passing through the working electrode is formed in the transverse direction. With this method, the usage amount of the sample entering the channel cannot be controlled, and it will flow out from the exhaust hole to cause pollution. Another way of composition, as mentioned above, is the design of Winarta et al., that is, a middle layer containing a U-shaped opening is pasted on the upper end of the working electrode, and then a top cover with a vent hole in the middle is pasted on it, thus forming A liquid channel is formed from the liquid inlet to the middle of the vent. With this kind of structure, when the air vent is too small, there is still concern that the sample will flow out of the air vent; when the air vent is moderate, the sample will just stay on the edge of the air vent, especially in today's stress on the miniaturization of the test piece. When taking a slice, it is easy to cause contamination due to inadvertently touching the vent and causing the sample to flow out.

发明内容Contents of the invention

如上所述，要想减少检体的体积和同时快速分析，必须避免造成检体污染的可能性，需运用毛细与虹吸原理，并配合电极的设计才能达到上述的目的。本发明即是依据上述电极区的设计，得到一快速导流的电极反应区，所以本发明提供既快速又精确的电化学生物检测器，结构简单容易生产且完全不需覆盖网膜，并且可避免检体外漏造成污染。As mentioned above, in order to reduce the volume of the sample and analyze quickly at the same time, the possibility of contamination of the sample must be avoided, and the above-mentioned purpose must be achieved by using the principle of capillary and siphon and cooperating with the design of the electrode. The present invention is based on the design of the above-mentioned electrode area to obtain a fast-conducting electrode reaction area, so the present invention provides a fast and accurate electrochemical biodetector, which is simple in structure and easy to produce and does not need to cover the omentum at all, and can Avoid contamination caused by external leakage of the specimen.

本发明的目的在于提供一种电化学式生物感测试片，其结合毛细虹吸原理的设计，可达到使用极少量检体既可快速又精确地进行电化学生物检测的目的。The object of the present invention is to provide an electrochemical biosensing test strip, which is designed in combination with the principle of capillary siphon, so as to achieve the purpose of fast and accurate electrochemical biodetection with a very small amount of sample.

本发明的目的还在于提供一种电化学式生物感测试片的制法，得到的试片进行生物检测既快速又精确，结构简单并且可避免检体外漏造成污染。The object of the present invention is also to provide a preparation method of an electrochemical biosensing test piece, the obtained test piece is fast and accurate for biological detection, has a simple structure and can avoid contamination caused by external leakage of the test piece.

本发明提供的电化学式生物感测试片，由下述组件构成：The electrochemical biosensing test piece provided by the present invention is composed of the following components:

一个绝缘底板；an insulating base plate;

一导线层，其是以网版印刷方式于绝缘底板上刷制而成的具有双电极式或三电极式导线；A wire layer, which is made by screen printing on the insulating base plate and has double-electrode or three-electrode wires;

一电极层，其是以网版印刷方式覆盖于导线层上，以形成含有一工作电极及一参考电极的双电极式、或含有一工作电极、一参考电极及一辅助电极的三电极式，该电极层的一端为电极区，另一端为与检测仪相连的接点，其中除参考电极区外，导线层完全被电极层所覆盖；An electrode layer, which is covered on the wire layer by screen printing to form a double-electrode type containing a working electrode and a reference electrode, or a three-electrode type containing a working electrode, a reference electrode and an auxiliary electrode, One end of the electrode layer is the electrode area, and the other end is the contact point connected to the detector, wherein except for the reference electrode area, the wire layer is completely covered by the electrode layer;

电绝缘中间层，其是以绝缘胶直接刷制或以绝缘材料直接粘贴覆盖于电极层上，使露出所述电极区及接点部分，该电绝缘中间层在相对于所述电极区的部分具有一U型或T型孔道；The electrical insulating intermediate layer is directly brushed with insulating glue or directly pasted and covered on the electrode layer with insulating material, so that the electrode area and the contact part are exposed, and the electrical insulating intermediate layer has a A U-shaped or T-shaped channel;

反应活性层，其含有缓冲溶液、填充物、反应催化物、电子媒介物及表面活性剂等物质，且在前述U型或T型孔道中涂覆于电极层表面；Reactive layer, which contains buffer solution, filler, reaction catalyst, electronic medium, surfactant and other substances, and is coated on the surface of the electrode layer in the aforementioned U-shaped or T-shaped channel;

上盖，其具有一相对于电绝缘中间层向上凸起的密闭空间，该上盖贴覆于电绝缘中间层上，使上盖与前述U型或T型孔道之间形成一毛细导流区，所述向上凸起的密闭空间对应U型孔道的内侧端底或T型孔道的纵横线交点处。The upper cover has a closed space that protrudes upwards relative to the electrical insulating intermediate layer, and the upper cover is attached to the electrical insulating intermediate layer, so that a capillary conduction area is formed between the upper cover and the aforementioned U-shaped or T-shaped channel , the upwardly protruding closed space corresponds to the inner end bottom of the U-shaped channel or the intersection of the vertical and horizontal lines of the T-shaped channel.

本发明还提供了一种所述电化学式生物感测试片的制法，步骤如下：The present invention also provides a method for preparing the electrochemical biosensing test piece, the steps are as follows:

a、以网版印刷技术在绝缘底板的一面上，印刷出一导线层，然后在40℃至120℃之下烘干；其中所述导线层为双电极式或三电极式；a. Print a wire layer on one side of the insulating base plate with screen printing technology, and then dry it at 40°C to 120°C; wherein the wire layer is a double-electrode type or a three-electrode type;

b、以网版印刷技术在前述的导线层上刷上电极层，再于40℃至120℃的下烘干；其中所述电极层为含有一工作电极及一参考电极的双电极式、或含有一工作电极、一参考电极及一辅助电极的三电极式，且此电极层的一端为电极区，另一端为与检测仪相连的接点，其中除参考电极区外，导线层完全被电极层所覆盖；b. Brush the electrode layer on the above-mentioned wire layer by screen printing technology, and then dry it at 40°C to 120°C; wherein the electrode layer is a double-electrode type containing a working electrode and a reference electrode, or A three-electrode type that contains a working electrode, a reference electrode and an auxiliary electrode, and one end of the electrode layer is the electrode area, and the other end is the contact point connected to the detector. Except for the reference electrode area, the wire layer is completely covered by the electrode layer. covered by;

c、利用绝缘胶层在前述印有电极层的绝缘底板上设置一含有U型或T型孔道的电绝缘中间层，使电极层一端的电极区位于U型或T型孔道内，并使电极层另一端的接点露出；c. Use an insulating adhesive layer to set an electrically insulating intermediate layer containing a U-shaped or T-shaped hole on the aforementioned insulating base plate printed with an electrode layer, so that the electrode area at one end of the electrode layer is located in the U-shaped or T-shaped hole, and make the electrode The contacts at the other end of the layer are exposed;

d、在所述U型或T型孔道中涂覆一反应活性层；d. Coating a reactive layer in the U-shaped or T-shaped channel;

e、于电绝缘中间层上粘贴一上盖，该上盖具有一相对于电绝缘中间层向上凸起的密闭空间，且该向上凸起的密闭空间位于所述U型孔道的内侧端底或T型孔道的纵横线交叉处，并使该上盖与U型或T型孔道之间形成一毛细导流区。e. Paste an upper cover on the electrical insulating intermediate layer, the upper cover has a closed space protruding upward relative to the electrical insulating intermediate layer, and the upwardly protruding enclosed space is located at the inner end bottom of the U-shaped channel or The intersection of the vertical and horizontal lines of the T-shaped channel forms a capillary diversion area between the upper cover and the U-shaped or T-shaped channel.

本发明依据毛细与虹吸结合原理以及电极区的设计，得到一快速导流的电极反应区，其仅需要极少的检体(约0.5-0.8μl)就可让电极正常工作，全部检测时间约为5-10秒，该试片是一个既快速又精确的电化学生物检测器，结构简单，完全不需覆盖网膜，并且可避免检体外漏造成污染。Based on the combination principle of capillary and siphon and the design of the electrode area, the present invention obtains a fast-conducting electrode reaction area, which requires only a small sample (about 0.5-0.8 μl) to make the electrode work normally, and the total detection time is about It takes 5-10 seconds. The test piece is a fast and accurate electrochemical biodetector with a simple structure, no need to cover the omentum at all, and can avoid contamination caused by external leakage of the specimen.

上述的绝缘底板是聚碳酸酯、聚酯、聚醚、聚酰胺、聚氨酯、聚亚胺、聚氯乙烯、玻璃、玻璃纤维板、陶瓷、聚丙烯及聚乙烯中的任一材料；导线层(导电层)是银、氯化银或金中的任何一种材料；电极层是碳、银、金或铂中的任何一种材料；反应催化物是生物性催化物如酵素或其它非生物性催化物。The above-mentioned insulating bottom plate is any material in polycarbonate, polyester, polyether, polyamide, polyurethane, polyimide, polyvinyl chloride, glass, glass fiber board, pottery, polypropylene and polyethylene; layer) is any material in silver, silver chloride or gold; the electrode layer is any material in carbon, silver, gold or platinum; the reaction catalyst is a biological catalyst such as enzyme or other non-biological catalysis thing.

上述的电绝缘中间层厚度介于20-400微米，最优选厚度为50-200微米；该电绝缘中间层的缺口孔道长度介于2-8毫米，宽度介于0.5-5毫米之间；相对于电绝缘中间层向上凸起的密闭空间体积介于0.5-4微升。The thickness of the above-mentioned electrical insulating interlayer is between 20-400 microns, and the most preferred thickness is 50-200 microns; the length of the gap channel of the electrical insulating interlayer is between 2-8 millimeters, and the width is between 0.5-5 millimeters; relatively The volume of the enclosed space protruding upward from the electrically insulating interlayer ranges from 0.5 to 4 microliters.

上述中间层的制法可以直接粘贴绝缘中间层或以网版印刷方式；该中间层的缺口孔道可以是U型或T型，该T型的横向缺口为向试片两侧的排气口。The method of making the above-mentioned intermediate layer can be pasting the insulating intermediate layer directly or by screen printing; the notch channel of the intermediate layer can be U-shaped or T-shaped, and the transverse notch of the T-shaped is an air outlet to both sides of the test piece.

该试片具有快速吸收检体、有效控制检测检体体积以及检体填充定位的特性。该试片的结构试在一绝缘底板上具有两个或三个电极，分别为工作电极、辅助电极及参考电极(三极式)；在电极区表面直接吸附一层反应层；反应层是由缓冲溶液、反应催化物、电子传递物质、润湿剂与表面活性剂等组成；电极区上方则有一检体导流区，分别由于电极区上方具有一U型通道的中间层薄片以及其上的上盖片组合而成.所形成的中空沟槽具有毛细与虹吸现象，能迅速将极少量的检体吸入至电极反应区，然后利用电化学的定电位法或定电流法，测定检体与反应区的反应催化物和电子传递物质所产生的反应，进而可以分析定量检体中的标的成分。上盖片上于电极区临导线前端具有一密闭凸状空间，具有有效控制检体体积以及检体填充定位的功能。The test piece has the characteristics of rapidly absorbing the specimen, effectively controlling the volume of the specimen to be tested, and filling and positioning the specimen. The structure of the test piece has two or three electrodes on an insulating base plate, which are respectively working electrode, auxiliary electrode and reference electrode (three-pole type); a layer of reaction layer is directly adsorbed on the surface of the electrode area; the reaction layer is composed of buffer solution, reaction catalysts, electron transfer substances, wetting agents and surfactants; above the electrode area, there is a specimen diversion area, respectively due to the middle layer sheet with a U-shaped channel above the electrode area and the The upper cover is combined. The formed hollow groove has the phenomenon of capillary and siphon, which can quickly suck a very small amount of sample into the electrode reaction area, and then use the electrochemical constant potential method or constant current method to measure the sample and The reaction produced by the reaction catalyzer in the reaction zone and the electron transfer substance can then analyze the target components in the quantitative sample. There is a closed convex space on the upper cover sheet adjacent to the front end of the wire in the electrode area, which has the function of effectively controlling the volume of the sample and filling and positioning the sample.

附图说明Description of drawings

图1：本发明的网版印刷电极试片具U型孔洞结构示意图；Fig. 1: Schematic diagram of the U-shaped hole structure of the screen printing electrode test piece of the present invention;

图2：本发明的网版印刷电极试片具T型孔洞结构示意图；Fig. 2: Schematic diagram of the T-shaped hole structure of the screen printing electrode test piece of the present invention;

图3：本发明的网版印刷电极试片纵切面示意图；Fig. 3: schematic diagram of longitudinal section of screen printing electrode test piece of the present invention;

图4：本发明的内含式向上突起密闭式空间上盖结构示意图；Figure 4: Schematic diagram of the structure of the built-in upwardly protruding closed space upper cover of the present invention;

图5：本发明的内含式向上突起密闭式空间上盖结构纵切面示意图；Figure 5: Schematic diagram of the longitudinal section of the built-in upwardly protruding closed space upper cover structure of the present invention;

图6：本发明的内含式向上突起密闭式空间网版印刷电极试片纵切面结构示意图；Fig. 6: Schematic diagram of the structure of the vertical section of the electrode test piece of the built-in upwardly protruding airtight space screen printing electrode test piece of the present invention;

图7：使用本发明的电极试片时，全血检体体积对检测的影响。Figure 7: The effect of whole blood sample volume on detection when using the electrode test strip of the present invention.

具体实施方式Detailed ways

以下结合具体实施例详细说明本发明，但不限定本发明的实施范围。The present invention will be described in detail below in conjunction with specific examples, but the implementation scope of the present invention is not limited.

感测试片Sensory test piece

本发明的电化学三极式网版印刷生物感测试片结构如图1所示，在绝缘性底板1，如聚氯乙烯(PVC)、聚酯(PE)、聚碳酸酯(PC)等，上面以导电性材料，例如银、金等胶状材料，以网版印刷方式刷制成导线2，再以导电性胶状材料如碳、金或白金再印刷覆盖于导线2之上，形成电极的长条，其一端如工作电极3、参考电极4、辅助电极5(两极式设计时则无参考电极)；另一端如3’、4’与5’于相对电极的端点则为与检测仪的相接点，而6为侦测仪的试片自动辨识线。在印有电极的底板上粘贴或以网版印刷方式刷上一含有U型孔洞的不导电中间层或绝缘胶层7，其为绝缘介电层兼具间隔功能。沟槽7a为检体导流槽，上盖8于相对导流槽底部形成一个密闭且向上突起的空间8a，其内部体积约2μl。电极反应区涂覆试剂配方，包括了缓冲溶液、反应催化物(例如酵素)、电子传导物质(例如dimethyl ferrocene、tetrathiofulvalene等)、润湿剂(纤维素cellulose、羟乙基纤维素hydroxyethyl cellulose、聚乙烯醇polyvinyl alcohol、乙烯聚合物polyvinyl、吡咯烷酮pyrrolidone、明胶gelatin等)及表面活性剂(tween20、triton X-100、surfynol、mega8等)。间隔层7与上盖层8贴合后在电极工作区周围形成毛细导流通道7a，允许检体如血液由7a的前缘接触后，迅速经由毛细现象填满至电极反应区，此时由检体与反应物经由反应催化物催化和电子传递物质作用，再经由电极及检测仪侦测产生电流的讯号。上述的导流沟槽可以在非常快速(小于1秒)且需要很少血液(小于1微升)的情形下，让电极进行正常的侦测工作.The structure of the electrochemical three-pole screen-printed biosensing test piece of the present invention is as shown in Figure 1, on the insulating base plate 1, such as polyvinyl chloride (PVC), polyester (PE), polycarbonate (PC) etc., Conductive materials, such as silver, gold and other colloidal materials, are used to screen-print theconductive wire 2, and then printed with conductive colloidal materials such as carbon, gold or platinum to cover theconductive wire 2 to form electrodes. One end of which is the workingelectrode 3,reference electrode 4, auxiliary electrode 5 (there is no reference electrode in the bipolar design); the other end is 3', 4' and 5' at the end of the opposite electrode, which is connected to the detector , and 6 is the automatic identification line of the test piece of the detector. A non-conductive intermediate layer or an insulating adhesive layer 7 containing U-shaped holes is pasted or screen-printed on the bottom plate printed with electrodes, which is an insulating dielectric layer and has a spacer function. Thegroove 7a is the sample diversion groove, and theupper cover 8 forms a closed and upwardlyprotruding space 8a opposite to the bottom of the diversion groove, and its internal volume is about 2 μl. The electrode reaction area coating reagent formula includes a buffer solution, a reaction catalyst (such as an enzyme), an electron-conducting substance (such as dimethyl ferrocene, tetrathiofulvalene, etc.), a wetting agent (cellulose cellulose, hydroxyethyl cellulose, poly Vinyl alcohol polyvinyl alcohol, ethylene polymer polyvinyl, pyrrolidone pyrrolidone, gelatin gelatin, etc.) and surfactants (tween20, triton X-100, surfynol, mega8, etc.). After the spacer layer 7 and theupper cover layer 8 are bonded together, acapillary flow channel 7a is formed around the electrode working area, which allows the sample such as blood to be quickly filled to the electrode reaction area through capillary phenomenon after being contacted by the front edge of 7a. The sample and the reactant are catalyzed by the reaction catalyst and acted on by the electron transfer material, and then the signal of the generated current is detected by the electrode and the detector. The above-mentioned diversion groove can allow the electrodes to perform normal detection work very quickly (less than 1 second) and requires very little blood (less than 1 microliter).

本发明另一电化学三极式电极试片结构如图2所示。简述之，在绝缘底板1上以导电材料如银、氯化银、金等材料以网版印刷方式刷上导电层2，再以导电性材料如碳、金或白金等刷制覆盖于导电层上形成工作电极3、参考电极4及辅助电极5。其各电极相对的一端3’、4’及5’为与检测仪相接的接点；而6为侦测仪的试片自动辨识线。下一步骤为在完成电极层的底板上粘贴或以绝缘胶以网版印刷方式刷上一个T字型孔洞的间隔层7；上盖层8包含一密闭且向上凸起的空间8a，此凸起空间8a的内部体积约2μl，位于T字纵横线交点的正上方；而上盖与间隔层间形成的沟槽7a为检体导流槽，7b及7c为朝向两侧的排气通道。检体如血液等，由7a的前缘接触后，由毛细原理迅速流经填满至电极反应区，而检体的前缘亦如图1的设计，不超过8a的前端；另外，该设计可删除7b或7c任一侧的排气通道，亦可获得相同的效果。The structure of another electrochemical three-pole electrode test piece of the present invention is shown in FIG. 2 . Briefly, theconductive layer 2 is brushed on the insulating base plate 1 by screen printing with conductive materials such as silver, silver chloride, gold, etc., and then covered with conductive materials such as carbon, gold, or platinum. A workingelectrode 3 , areference electrode 4 and anauxiliary electrode 5 are formed on the layer. The opposite ends 3', 4' and 5' of each electrode are the contact points connected with the detector; and 6 is the automatic identification line of the test piece of the detector. The next step is to paste a spacer layer 7 with a T-shaped hole on the bottom plate of the electrode layer or use insulating glue to screen-print; theupper cover layer 8 includes a closed and upwardly raisedspace 8a, which The internal volume of thespace 8a is about 2 μl, and it is located directly above the intersection of the vertical and horizontal lines of the T; thegroove 7a formed between the upper cover and the spacer layer is the sample diversion groove, and 7b and 7c are exhaust channels facing both sides. Specimens, such as blood, etc., after being contacted by the leading edge of 7a, quickly flow through and fill up to the electrode reaction area by capillary principle, and the leading edge of the specimen is also designed as shown in Figure 1, which does not exceed the front end of 8a; in addition, this design The exhaust channel on either side of 7b or 7c can be deleted to obtain the same effect.

底板基材Backplane base material

底板1可以由各种绝缘性材料组成，例如高分子、塑料以及陶瓷材料等，选用材料的种类必须配合电极材料以及应用上的需求，如果是侵入式用途则应选用软性材料以减轻疼痛并避免伤及组织，此类用途通常可以使用绝缘性高分子材料例如聚碳酸酯、聚酯(例如杜邦公司的Mylar系列产品)、聚乙烯对苯二酸盐(polyethylene terephthalate，PET)、聚氯乙烯(PVC)、聚醚、聚酰胺、聚氨酯、聚亚胺等；另一方面刚性材料可使用不易断裂或弯曲者如陶瓷材料像二氧化硅或二氧化铝等。以体外检测用途而言，底板的尺寸宽度一般通常介于3-15毫米，更精确的可介于5-10毫米，厚度约在50-800微米或更精确的介于200-400微米，底板长度则受各种因素影响可能介于1-8厘米，更精确的介于2-5厘米。Bottom plate 1 can be made of various insulating materials, such as polymers, plastics, and ceramic materials. The type of material selected must match the electrode material and application requirements. For invasive applications, soft materials should be used to relieve pain and prevent To avoid damage to tissues, insulating polymer materials such as polycarbonate, polyester (such as DuPont's Mylar series products), polyethylene terephthalate (polyethylene terephthalate, PET), polyvinyl chloride can usually be used for such purposes. (PVC), polyether, polyamide, polyurethane, polyimide, etc.; on the other hand, rigid materials can use those that are not easy to break or bend, such as ceramic materials such as silicon dioxide or aluminum dioxide. For in vitro detection purposes, the size and width of the bottom plate is generally between 3-15 mm, more precisely 5-10 mm, and the thickness is about 50-800 microns or more precisely between 200-400 microns. The length is affected by various factors and may be between 1-8 cm, more precisely between 2-5 cm.

导线层与电极Wire layer and electrode

如图1所示，导电性材料如银、金、白金等材料经网版印刷方式在底板1上形成导线区2，作为连接各电极与测试装置的用途，使用高导电性与低电阻特性的材料可以降低电极的阻抗，提高侦测电流讯号。导电性材料如碳胶则刷在导线2之上，6作为侦测仪的试片自动辨识用途，除了参考电极4外，所刷制的导线2被完全覆盖，4的外露银导线部分利用电化学原理可以再将银表面处理成氯化银的拟参考电极，或者以银/氯化银油墨刷制覆盖于银导线上，或直接以银/氯化银油墨刷制导线，这样就不需另外作氯化银处理。As shown in Figure 1, conductive materials such as silver, gold, platinum and other materials are screen-printed to form awire area 2 on the bottom plate 1. As the purpose of connecting each electrode and the test device, awire area 2 with high conductivity and low resistance characteristics is used. The material can reduce the impedance of the electrode and improve the detection current signal. Conductive material such as carbon glue is brushed on thewire 2, and 6 is used for the automatic identification of the test piece of the detector. Except for thereference electrode 4, the brushedwire 2 is completely covered, and the exposed silver wire part of 4 is covered with an electric wire. The chemical principle can then treat the silver surface into a quasi-reference electrode of silver chloride, or cover the silver wire with silver/silver chloride ink, or directly brush the wire with silver/silver chloride ink, so that no need In addition, it is treated with silver chloride.

绝缘层Insulation

绝缘中间层7，可由具绝缘性的介电材料刷制而成，或以粘贴方式覆盖于电极表面，除了遮蔽不必外露的碳表面外，还提供一固定面积的反应区.The insulating interlayer 7 can be brushed from an insulating dielectric material, or covered on the electrode surface by pasting. In addition to covering the carbon surface that does not need to be exposed, it also provides a fixed area of the reaction area.

反应试剂区Reagent area

反应试剂涂覆在电极区之上，包括了反应用的反应催化物、缓冲溶液、填充物、电子传递物质、表面活性剂等。例如使用在葡萄糖检测时反应催化物可以是葡萄糖氧化酶或脱氢酶，填充物成分包括了高分子或湿润剂例如纤维素、聚乙醇、明胶等，表面活性剂成分，例如Tween-80、Triton X-100、Surfynol或Mega 8等等，提供检体与试剂回溶分散功能，以及毛细导流区的亲水性与分散功能。因此，反应试剂层可同时提供反应与毛细功能，不但让检体能充满电极区并使分析反应进行，提供电极反应电流作为定量检体成分的功能。较适合的电子传递物质应具有介于-100至+500毫伏的氧化还原电位，依不同的检测需求而应用不同的电子传递物质.例如检测葡萄糖时可应用二茂铁系列的二甲基二茂铁(dimethyl ferrocene)以及四硫富瓦烯(tetrathiafulvalene)或上述二者的衍生物或错合物，较低的电位可以避免检体中的干扰物质，较高的电子传递效率则可提供较强的电流讯号。缓冲溶液作用在于维持一定范围的酸碱值，一般酸碱值介于PH4-9，较适合酸碱值介于PH5-8之间.可以使用的缓冲液成分包括磷酸盐、醋酸盐、柠檬酸盐等，浓度范围介于10-1000毫摩尔浓度，最适浓度的30-1000毫摩尔浓度。The reaction reagents are coated on the electrode area, including reaction catalysts, buffer solutions, fillers, electron transfer substances, surfactants, etc. for the reaction. For example, when using glucose detection, the reaction catalyst can be glucose oxidase or dehydrogenase, and the filler components include polymers or wetting agents such as cellulose, polyethylene glycol, gelatin, etc., surfactant components, such as Tween-80, Triton X-100, Surfynol orMega 8, etc., provide the function of sample and reagent resolubility and dispersion, as well as the hydrophilicity and dispersion function of the capillary diversion area. Therefore, the reaction reagent layer can provide reaction and capillary functions at the same time, not only allowing the sample to fill the electrode area and allowing the analysis reaction to proceed, but also providing the electrode reaction current as a function of quantifying the sample components. A more suitable electron transfer substance should have an oxidation-reduction potential between -100 and +500 millivolts, and different electron transfer substances should be used according to different detection requirements. For example, the ferrocene series of dimethyl di Ferrocene (dimethyl ferrocene) and tetrathiafulvalene (tetrathiafulvalene) or derivatives or complexes of the above two, lower potential can avoid interfering substances in the sample, and higher electron transfer efficiency can provide better Strong current signal. The function of the buffer solution is to maintain a certain range of pH value. Generally, the pH value is between PH4-9, and the more suitable pH value is between PH5-8. The buffer components that can be used include phosphate, acetate, lemon salt, etc., the concentration range is between 10-1000 millimolar concentration, and the optimum concentration is 30-1000 millimolar concentration.

毛细导流层Capillary drainage layer

导流层由间隔薄片7与上盖8覆盖于电极上方所形成；7a为检体毛细沟槽，如具有向两侧排气功能(T字型设计)则7b与7c为排气口，7b或7c可单独存在。利用薄片7的厚度及7a沟槽的宽度可以调整导流区体积的大小，一般来说厚度介于20-400微米，优选厚度50-200微米，中空区长度介于2-8毫米，宽度介于0.5-5毫米，优选宽度则介于1-2毫米，所形成的中空区域体积介于0.2-160微升，实际检体操作时最适量约介于0.5-2微升，检体接触导流层边缘时到充满所需时间小于1秒钟。The diversion layer is formed by covering the electrode with the spacer sheet 7 and theupper cover 8; 7a is the capillary groove of the specimen, if it has the function of exhausting to both sides (T-shaped design), 7b and 7c are the exhaust ports, 7b Or 7c can exist alone. Utilize the thickness of sheet 7 and the width of 7a groove can adjust the size of diversion area volume, generally speaking thickness is between 20-400 micron, preferred thickness is 50-200 micron, and the length of hollow area is between 2-8 millimeter, and width is between In 0.5-5 mm, the preferred width is 1-2 mm, the volume of the formed hollow area is 0.2-160 microliters, the optimal amount is about 0.5-2 microliters during actual sample operation, and the sample contact conduction The time required from the edge of the flow layer to full is less than 1 second.

上盖层8的封闭式凸起8a可以是圆形、方形或其它几何形状，适当的大小介于0.5-4微升，缺口的位置位于工作电极导流通道的后上方，此缺口能使进入的血液检体填满反应区后停止前进。间隔片7与上盖片8可以由透明或不透明绝缘性材料组成，例如塑料或高分子材料如PVC聚氯乙烯或Mylar等，导流区上方区域的8a可呈透明状，除了使用者可目视检体是否顺利填满，还具有保护试片的功能。上盖层可以藉由两步骤形成。第一步骤是将图一所谓的上盖层的上端开一孔洞，亦即8a，再于此片上覆盖一完整的薄片9(如图4及图5所示)，图3及图6所显示图1及含薄片9试片的纵切面。Theclosed protrusion 8a of theupper cover layer 8 can be circular, square or other geometric shapes, and the appropriate size is between 0.5-4 microliters. Stop advancing after the blood sample fills the reaction zone. The spacer 7 and theupper cover 8 can be made of transparent or opaque insulating materials, such as plastics or polymer materials such as PVC polyvinyl chloride or Mylar, etc., thearea 8a above the diversion area can be transparent, except that the user can see Check whether the object is filled smoothly, and also has the function of protecting the test piece. The capping layer can be formed by two steps. The first step is to open a hole in the upper end of the so-called upper cover layer in Fig. 1, namely 8a, and then cover a complete thin sheet 9 (as shown in Fig. 4 and Fig. 5) on this sheet, as shown in Fig. 3 and Fig. 6 Fig. 1 and the longitudinal section of the testpiece containing slice 9.

注满传感器Fill sensor

注满传感器的设计用来检知三组电极上方是否充满检体，在三极式设计中若工作电极在导流区最外侧时，利用辅助电极与工作电极可完成注满感测功能，运用电流、电位或电阻值的监控，当检体尚未填满三个电极时，定电位监测工作与辅助电极间的阻抗值应为无限大，而当检体填满电极时，阻抗会急剧减小，经由此确定检体填满时才启动电化学部分的分析参数，在两电极设计时亦可利用同原理监控。利用电极作为注满传感器，必须使电极的分布与检体流动方向一致，也就是说工作电极最先接触检体，辅助电极必须最后接触检体方能正确判知是否填满，同样也可将辅助电极排在最先接触点。The filling sensor is designed to detect whether the sample is filled above the three sets of electrodes. In the three-electrode design, if the working electrode is on the outermost side of the diversion area, the auxiliary electrode and the working electrode can be used to complete the filling sensing function. For the monitoring of current, potential or resistance value, when the sample has not filled the three electrodes, the impedance value between the constant potential monitoring work and the auxiliary electrode should be infinite, and when the sample is filled with electrodes, the impedance will decrease sharply Through this, it is determined that the analytical parameters of the electrochemical part are started only when the sample is full, and the same principle can also be used for monitoring in the design of two electrodes. When using the electrode as a filling sensor, the distribution of the electrodes must be consistent with the flow direction of the sample, that is to say, the working electrode must first contact the sample, and the auxiliary electrode must contact the sample last to correctly judge whether it is filled. Auxiliary electrodes are arranged at the first point of contact.

电化学分析Electrochemical analysis

当电极组合完成经由分切或冲压方式裁切后，利用外接的掌上型电化学仪即可进行检体分析，分析方式可以多种方式进行，例如定电位法(0-0.6伏特)量测稳态电流，或者以定电位量测一定时间内的总电量值，电量值为电流与时间的积分，与稳态电流均可作为检体中成分浓度的正比检量关系，检测仪中配合电极片的注满感测功能，当检测仪读知注满讯号时自动启动电化学分析参数，如此增加检测的准确性，尤其当整体检测时间已低于10秒时，少许时间的误差往往会造成很大的变异。After the electrode combination is cut by slitting or punching, the sample can be analyzed using an external handheld electrochemical instrument. The analysis method can be carried out in various ways, such as the constant potential method (0-0.6 volts) to measure the stability of the sample. State current, or measure the total electricity value within a certain period of time with a constant potential. The electricity value is the integral of current and time, and the steady-state current can be used as a proportional measurement relationship of the component concentration in the sample. The detector is equipped with an electrode sheet The full-fill sensing function, when the detector reads the full-fill signal, automatically starts the electrochemical analysis parameters, thus increasing the detection accuracy, especially when the overall detection time is less than 10 seconds, a small time error will often cause a large big variation.

以下的实施例乃以血糖检测为例，其目的只是在阐述本发明的可实行性而并不对本发明范围有所局限。The following embodiments take blood glucose detection as an example, and the purpose is only to illustrate the practicability of the present invention without limiting the scope of the present invention.

实施例一、网版印刷葡萄糖检测试片制作Embodiment 1, Screen Printing Glucose Detection Test Strip Production

聚丙烯合成纸底板以300网目的网版印刷导电银胶层，在50℃下干燥30分钟后，再以碳胶刷上三组电极(工作电极、参考电极及辅助电极)。再次于90℃烘烤15分钟，然后以绝缘胶刷制后于紫外光下硬化干燥形成绝缘层，并露出检导流反应区7a、7b以及7c(有通气孔的设计)。反应试剂2-6微升，(内含0.5-3单位葡萄糖氧化酶、0.1-1％聚乙烯醇、pH4.0-9.0，100mM磷酸钾缓冲液、10-100mM氯化钾、二甲基二茂铁0.05-0.5％、tween-20The polypropylene synthetic paper bottom plate is printed with a conductive silver glue layer with a 300-mesh screen, dried at 50°C for 30 minutes, and then brushed with three sets of electrodes (working electrode, reference electrode and auxiliary electrode) with carbon glue. Bake again at 90° C. for 15 minutes, then brush with insulating glue, harden and dry under ultraviolet light to form an insulating layer, and expose the flow detection andreaction regions 7 a , 7 b and 7 c (designed with vent holes). Reagent 2-6 microliters, (containing 0.5-3 units of glucose oxidase, 0.1-1% polyvinyl alcohol, pH4.0-9.0, 100mM potassium phosphate buffer, 10-100mM potassium chloride, dimethyl di Ferrocene 0.05-0.5%, tween-20

0.005-0.2％、surfynol0.005-0.2％及碳酸甲基纤维0.1％-1.0％)，涂覆于前述所形成的凹陷导流区7a，于45℃干燥1小时后取出再贴合含有中央孔洞的上盖片8，粘贴后再以一透明上盖片9压合固着后再分切即为完成品。0.005-0.2%, surfynol 0.005-0.2%, and methyl carbonate fiber 0.1%-1.0%), coated on theconcave diversion area 7a formed above, dried at 45°C for 1 hour, then taken out and pasted with a central hole Theupper cover sheet 8 is pasted and then cut with a transparentupper cover sheet 9 after being pressed and fixed to be a finished product.

实施例二、标准葡萄糖溶液与全血测试Embodiment two, standard glucose solution and whole blood test

配置标准磷酸钾缓冲溶液(PH7.4)内含葡萄糖浓度0-400毫克/100毫升，以实施例一所制备而成的网版印刷电极葡萄糖试片，在电化学分析仪(CHInstrument Co.650A)于测试电位100毫伏的状况下，测试时间为8秒钟，每次测试时，供应检体体积为3微升，每次试片吸入的体积皆小于3微升。其测试结果如表1所示.Configure standard potassium phosphate buffer solution (PH7.4) to contain glucose concentration 0-400 mg/100 milliliters, with the screen printing electrode glucose test piece prepared by embodiment one, in electrochemical analyzer (CHInstrument Co.650A ) under the condition of a test potential of 100 millivolts, the test time is 8 seconds, the volume of the supplied sample is 3 microliters for each test, and the volume inhaled by each test piece is less than 3 microliters. The test results are shown in Table 1.

表1 标准葡萄糖测试结果Table 1 Standard Glucose Test Results

 葡萄糖浓度(mg/dl)电压(微库仑)00.690251.532502.952 Glucose concentration (mg/dl) Voltage (microcoulomb) 0 0.690 25 1.532 50 2.952

 1005.2482007.4004009.577 100 5.248 200 7.400 400 9.577

全血检体亦可由本试片测量。新鲜采得的静脉全血以标准葡萄糖添加所得检量，数据如表2所示，测试电位100毫伏特，提供测试全血体积为2微升。Whole blood samples can also be measured by this test piece. The freshly collected venous whole blood was added with standard glucose, and the data are shown in Table 2. The test potential was 100 millivolts, and the volume of whole blood for testing was 2 microliters.

表2 不同葡萄糖含量的全血检测结果Table 2 Whole blood test results with different glucose contents

 葡萄糖浓度(mg/dl)电量(微库仑)801.5561052.6361303.4401805.9462809.70738011.73348012.46458013.945 Glucose concentration (mg/dl) Power (microcoulomb) 80 1.556 105 2.636 130 3.440 180 5.946 280 9.707 380 11.733 480 12.464 580 13.945

实施例三、不同全血量的血糖检测Embodiment 3. Blood glucose detection of different whole blood volumes

利用实施例一所制成的电极试片，供应不同体积的全血测试的本发明的需求血量，静脉全血以标准葡萄糖液配制成300mg/dl的浓度。The electrode test strip made in Example 1 was used to supply different volumes of whole blood to test the blood volume required by the present invention, and the venous whole blood was prepared with a standard glucose solution to a concentration of 300 mg/dl.

本测试的方法为提供不同体积的全血，使试片以虹吸的原理吸入，并以如实施例二的检测条件进行检测。如图7结果显示，当吸入检体量不足时(如0.5μl)，检测葡萄糖浓度偏低；但当所提供吸入的检体体积达0.8μl以上时，葡萄糖检出浓度则趋近于所配制的浓度，且所提供的全血检体亦无法全部被吸入；所提供的检体体积愈高，剩余的体积则愈多，此表示检体导流反应区已达饱和，无法吸入更多的检体。所吸入检体的前缘皆不超过8a与导流区相接之处，足以证明本发明的设计有限制检体体积的功能。The method of this test is to provide different volumes of whole blood, so that the test piece is sucked in by the principle of siphon, and the test is carried out under the test conditions as in Example 2. The results shown in Figure 7 show that when the inhaled sample volume is insufficient (such as 0.5 μl), the detected glucose concentration is low; but when the provided inhaled sample volume reaches 0.8 μl or more, the detected glucose concentration tends to be close to the prepared one. concentration, and the provided whole blood sample cannot be inhaled completely; the higher the volume of the sample provided, the more the remaining volume, which means that the diversion reaction area of the sample is saturated, and no more sample can be inhaled. body. The front edge of the inhaled sample does not exceed the junction of 8a and the diversion area, which is enough to prove that the design of the present invention has the function of limiting the volume of the sample.

以上描述了本发明的优选实施例，然其并非用以限定本发明。本领域技术人员对在此公开的实施方案可进行并不偏离本发明范畴和精神的改进和变化。The preferred embodiments of the present invention are described above, but they are not intended to limit the present invention. Modifications and changes to the embodiments disclosed herein may be made by those skilled in the art without departing from the scope and spirit of the invention.

Claims (17)

1, a kind of electric chemical formula biological sensing test sheet is made of following assembly:

An insulating base;

-conductor layer, it is to have bipolar electrode formula or three electric pole type leads with what the screen painting mode system of brushing on insulating base formed;

One electrode layer, it is to be covered on the conductor layer in the screen painting mode, contain the bipolar electrode formula of a working electrode and a reference electrode or contain three electric pole types of a working electrode, a reference electrode and an auxiliary electrode with formation, one end of this electrode layer is an electrode district, the other end is the contact that links to each other with detector, wherein except that the reference electrode district, conductor layer is covered by electrode layer fully;

The electrical isolation middle layer, it is directly to paste with the insulating gel system of directly brushing or with insulating material to be covered on the electrode layer, makes and exposes described electrode district and contact portions, this electrical isolation middle layer has a U type or T type duct in the part with respect to described electrode district;

The reactivity layer, it contains buffer solution, filling material, catalytic reaction thing, electron mediator and surfactant, and is coated on electrode layer surface in aforementioned U type or T type duct;

Loam cake, it has a confined space that raises up with respect to the electrical isolation middle layer, this loam cake is covered on the electrical isolation middle layer, make and form a capillary water conservancy diversion district between loam cake and aforementioned U type or the T type duct, at the bottom of the medial extremity in the corresponding U type of the described confined space that raises up duct or the co-ordination intersection point place in T type duct.

2, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein insulating base is any material in polycarbonate, polyester, polyethers, polyamide, polyurethane, poly-imines, Polyvinylchloride, glass, glass mat, pottery, polypropylene and the tygon.

3, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein conductor layer is any material in silver, silver chloride or the gold.

4, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein electrode layer is a carbon, any material in silver, gold or the platinum.

5, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein the catalytic reaction thing is biological catalytic materials or inanimate catalytic materials.

6, electric chemical formula biological sensing test sheet as claimed in claim 5, wherein biological catalytic materials is a ferment.

7, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein the electrical isolation intermediate layer thickness is between the 20-400 micron.

8, electric chemical formula biological sensing test sheet as claimed in claim 7, wherein the electrical isolation intermediate layer thickness is the 50-200 micron.

9, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein the U type in electrical isolation middle layer or T type orifice throat length are between the 2-8 millimeter, and width is between the 0.5-5 millimeter.

10, electric chemical formula biological sensing test sheet as claimed in claim 1, wherein the confined space volume that raises up with respect to the electrical isolation middle layer of loam cake is between the 0.5-4 microlitre.

11, a kind of method for making of electric chemical formula biological sensing test sheet as claimed in claim 1, finish by following step:

A, with screen printing technology on the one side of insulating base, print out a conductor layer, then under 40 ℃ to 120 ℃ the oven dry; Wherein said conductor layer is bipolar electrode formula or three electric pole types;

B, on aforesaid conductor layer, brush electrode layer with screen printing technology, again in 40 ℃ to 120 ℃ following oven dry; Wherein said electrode layer is three electric pole types that contain the bipolar electrode formula of a working electrode and a reference electrode or contain a working electrode, a reference electrode and an auxiliary electrode, and an end of this electrode layer is an electrode district, the other end is the contact that links to each other with detector, wherein except that the reference electrode district, conductor layer is covered by electrode layer fully;

C, utilize the insulation glue-line that the one electrical isolation middle layer of containing U type or T type duct is set on the aforementioned insulating base that is printed on electrode layer, make the electrode district of electrode layer one end be positioned at U type or T type duct, and the contact of the electrode layer other end is exposed;

D, in described U type or T type duct, apply a reactivity layer;

E, on the electrical isolation middle layer, paste a loam cake, this loam cake has a confined space that raises up with respect to the electrical isolation middle layer, and this confined space that raises up is positioned at the bottom of the medial extremity in described U type duct or the co-ordination infall in T type duct, and makes and form a capillary water conservancy diversion district between this loam cake and U type or the T type duct.

12, method for making as claimed in claim 11, wherein the method for making in electrical isolation middle layer is to be provided with direct stickup insulating intermediate layer or in the screen painting mode.

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