WO2018068582A1 - 一种免疫层析检测装置 - Google Patents

一种免疫层析检测装置 Download PDF

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WO2018068582A1
WO2018068582A1 PCT/CN2017/098112 CN2017098112W WO2018068582A1 WO 2018068582 A1 WO2018068582 A1 WO 2018068582A1 CN 2017098112 W CN2017098112 W CN 2017098112W WO 2018068582 A1 WO2018068582 A1 WO 2018068582A1
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sample
test strip
detector
detection
fluorescent
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PCT/CN2017/098112
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English (en)
French (fr)
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宋旭东
黄若磐
徐部灼
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广州瑞博奥生物科技有限公司
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    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
    • 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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • G01N33/54386—Analytical elements
    • G01N33/54387—Immunochromatographic test strips
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence

Definitions

  • the present invention relates to the field of immunoassay technology, and in particular to an immunochromatographic detection device.
  • Immunochromatography is a rapid diagnostic technique developed in recent years. The principle is to fix a specific antibody to a certain zone of the nitrocellulose membrane. When the dried nitrocellulose is immersed in the sample. After (urine or serum), the sample will move forward along the membrane due to capillary action. When moving to the region where the antibody is immobilized, the corresponding antigen in the sample will specifically bind to the antibody, if immunocolloid is used. Gold or immunoenzymatic staining allows the region to display a certain color to provide qualitative or semi-quantitative detection of the sample for specific immunodiagnosis. Compared with the conventional diagnostic method, the technique is fast, and all the detection processes only take 5 to 30 minutes.
  • the immunochromatographic test strip is prepared by combining the reaction specificity and chromatographic technique of the immunolabel with the corresponding antigen (antibody).
  • the immunochromatographic test strip usually consists of a loading zone, a reaction zone and a water absorption zone.
  • the sample application zone contains immunolabel particles, which are usually adsorbed by the glass fibers in the region; the reaction zone is sprayed with two reaction lines, one for the detection line and one for the quality control line.
  • the detection line is used to detect the reactivity of the (antigen) substance here with the coating antigen (antibody or antibody-carrying antigen) of the immunolabel particle; the quality control line is used to detect the activity of the coated protein on the immunolabel particle.
  • the extent to which the immunolabel is released from the sample-adding zone passes through the reaction zone, and partially proceeds to the adsorption zone to complete the chromatography, thereby achieving rapid detection.
  • the latest immunochromatography uses chemical cross-linking or biomolecular specific interaction to couple specific antibodies to the surface of fluorescent molecules or fluorescent particles to obtain fluorescent dye-modified specific antibodies and fluorescent dye-modified quality control molecules, respectively.
  • a reactive group is present on the surface of the fluorescent dye, it can be directly reacted with a specific antibody without using a chemical crosslinking agent; otherwise, a fluorescent crosslinking agent is required to couple the fluorescent dye with a specific antibody or a quality control molecule.
  • the chemical crosslinking agent includes 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), N-hydroxysuccinimide (NHS), glutaraldehyde and the like.
  • the fluorescent dye In order to improve the discrimination between the signal and the background, the fluorescent dye should have a fluorescence wavelength range of 550 to 800 nm. Because under UV irradiation, the autofluorescence intensity of the chromatographic membrane, the bottom plate and the buckle will be much stronger than 550 nm below 550 nm, which will have a certain influence on the detection of low concentration protein. Therefore, it is preferred to emit a fluorescent dye with a wavelength greater than 550 nm. In addition, the chromatographic film, the substrate, and the card are generally weak in fluorescence intensity in the near-infrared region (600 to 800 nm). Therefore, a fluorescent dye having a wavelength in the range of 550 to 800 nm is preferable to further improve the sensitivity. Fluorescent dyes include organic fluorescent dyes and rare earth fluorescent dyes such as the AlexaFluro series of fluorescein.
  • the currently used immunofluorescence chromatography device uses a reflection method to detect the fluorescent signal on the porous membrane, and the fluorescence detector captures a specific antibody modified by the fluorescent dye on the surface of the porous membrane, and the fluorescent signal inside the porous membrane is not detected, resulting in detection. The sensitivity is reduced.
  • the present invention provides an immunochromatography detecting device which is creatively employed in a transmission method capable of capturing fluorescent signals on the surface and inside of a multi-layered porous film to make the detection result more accurate.
  • An immunochromatographic detecting device consisting of a test strip comprising a multilayer porous membrane and a support sheet, the test strip having a light transmittance of 10% or less at a wavelength of less than 450 nm and a wavelength of 500 nm or more
  • the material has a light transmittance of more than 90%;
  • the detector cavity internally includes an excitation light source, a detection area in which the test strip is placed, and a photodetector, and the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • the components of the detector of the present invention are identical in composition to the components of the conventional immunochromatographic detector, the only difference being that the excitation light source and the photodetector in the detector of the present invention are respectively located on both sides of the detection area; and the conventional detection The excitation source and photodetector in the device are located on the same side of the detection zone.
  • the difference is that the conventional detector detects the fluorescent signal on the multilayer porous film by the reflection method, and the present invention detects the fluorescent signal on the multilayer porous film by the transmission method.
  • the present invention improves the material of the support sheet in the conventional test strip, and has a light transmittance of 10% or less at a wavelength of less than 450 nm and a transmittance of 90% or more at a wavelength of 500 nm or more. s material. This material allows most of the visible light to pass through. In this way, the excitation light source can penetrate the surface and the inside of the multilayer porous membrane, and the photodetector can capture the fluorescent signals on the surface and inside of the multilayer porous membrane, so that the detection result is more accurate.
  • the support sheet may be colorless or colored according to the requirements of the support sheet for different wavelengths of light source.
  • Plastic supports can incorporate dyes and pigments to attract photons of a certain wavelength to meet the transmission requirements for light sources of different wavelengths.
  • the support sheet is a polyester, acryl, polycarbonate, polyacrylate, ethylene glycol modified polyethylene terephthalate or cellulose acetate butyrate material.
  • the support sheet is a polycarbonate material.
  • Polycarbonate is a kind of optical resin material with high refractive index. Its light transmittance is equivalent to inorganic glass, its refractive index is 1.59, its density is half of inorganic glass, and it can be freely formed into any shape with good biocompatibility. .
  • the present invention has the following beneficial effects:
  • the traditional immunofluorescence chromatography method is to add a certain amount of sample to the sample pad, move forward through the layer plate, and the labeling reagent dissolved on the binding pad reacts with the sample, and when the conjugate moves to the antigen detecting zone, the test is to be tested.
  • the specific reaction of the substance with the conjugate is trapped, and the conjugate is enriched on the detection line, and the conjugate content is proportional to the content of the sample in the sample.
  • the intensity of the fluorescence generated under the excitation light source is proportional to the content of the conjugate on the test strip.
  • the strength of the electrical signal is related to the number of fluorescent molecules, and the detector calculates the content of the analyte in the sample.
  • the advantage of the transmission method over conventional reflection methods is the high sensitivity. Because the traditional reflection method only collects the antibody signal on the surface of the membrane, and the antibody reaction signal inside the transmission membrane can be collected, the signal will be higher.
  • Figure 1 is a perspective view of the detector.
  • Figure 2 is a schematic view of the internal structure of the detector cavity.
  • Figure 3 is a schematic diagram showing the results of detecting the card slot.
  • Figure 4 shows the results of comparison with hospital examination data.
  • test methods used in the following examples are conventional methods unless otherwise specified; the materials, reagents and the like used are, if not specified, commercially available reagents and materials.
  • a fluorescent immunochromatography detecting device comprises a test strip comprising a multi-layer porous membrane and a support sheet, and a multi-layer porous membrane respectively being a sample mat, a binding mat, a nitrocellulose membrane and a water absorbing material Paper, a fluorescent labeling reagent is fixed on the bonding pad, the nitrocellulose membrane is provided with a detection line and a control line, and the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet to obtain a test paper. article.
  • the support sheet is a polycarbonate material.
  • the polycarbonate material has a light transmittance of 10% or less at a wavelength of less than 400 nm, and a light transmittance of 90% or more at a wavelength of 500 nm or more, and can ensure transmission of ultraviolet light, visible light, and infrared light.
  • the schematic diagram of the detector is shown in Figure 1.
  • the internal structure of the detector cavity is shown in Figure 2.
  • the detector comprises a housing and an excitation light source in the housing, a detection card slot for placing the test strip, a photodetector, and a microcontroller.
  • the excitation light source and the photodetector are respectively located on two sides of the detection card slot, and the excitation light source and the light detection are respectively
  • the device is connected to the microcontroller through a detection circuit.
  • the surface of the casing of the detector is provided with a display screen and a card slot opening.
  • the detection card slot can be extracted from the detector housing through the opening, and the detection card slot can be pulled or popped, and the detection card slot is
  • the back-shaped card slot of the middle opening puts the test strip on the test card slot. After the two ends of the test strip are fixed by rotating the platen or other device, the test line and the control line on the test strip are just falling. At the intermediate opening of the mouth-shaped card slot, the transmitted light that excites the light source through the test strip can be detected by the photodetector without any blockage.
  • the fluorescent immunochromatography detecting device of the embodiment is used by adding a certain amount of the sample to be tested to the sample pad, moving forward through the layer plate, and the labeling reagent dissolved on the bonding pad reacts with the sample, when combined
  • the object moves to the antigen detection line
  • the specific reaction of the analyte and the conjugate is trapped, and the conjugate is enriched on the detection line, and the content of the conjugate is proportional to the content of the sample in the sample.
  • the intensity of the fluorescence generated under the excitation light source is proportional to the content of the conjugate on the test strip.
  • the transmitted light on the test strip is converted into an electrical signal.
  • the strength of the electrical signal is related to the number of fluorescent molecules.
  • the electrical signal is transmitted to the microcontroller.
  • the microcontroller receives the current signal, performs conversion processing analysis, and calculates the sample to be tested.
  • the content of the analyte is displayed by the microcontroller through the display on the surface of the detector housing.
  • a fluorescent immunochromatography detecting device comprises a test strip and a detecting device
  • the test strip comprises a multilayer porous membrane and a supporting sheet
  • the multilayer porous membrane is a sample mat, a bonding mat, a nitrocellulose membrane and an absorbent paper, respectively a fluorescent labeling reagent is fixed on the bonding pad
  • the nitrocellulose membrane is provided with a detection line and a control line
  • the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet, thereby obtaining Go to the test strip.
  • the support piece is an acrylic material.
  • the acrylic material has a light transmittance of 10% or less at a wavelength of 400 nm, and a material having a light transmittance of 90% or more at a wavelength of 500 nm or more, and can ensure transmission of ultraviolet light, visible light, and infrared light.
  • the detector has the same component composition as that of Embodiment 1 or the detector and the fluorescent chromatography detector commonly used in the prior art, except that the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • a fluorescent immunochromatography detecting device comprises a test strip and a detecting device
  • the test strip comprises a multilayer porous membrane and a supporting sheet
  • the multilayer porous membrane is a sample mat, a bonding mat, a nitrocellulose membrane and an absorbent paper, respectively a fluorescent labeling reagent is fixed on the bonding pad
  • the nitrocellulose membrane is provided with a detection line and a control line
  • the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet, thereby obtaining a test strip.
  • the support sheet is a polyester material.
  • the polyester material has a light transmittance of less than 10 nm and a light transmittance of 10% or less, and a material having a light transmittance of 90% or more at a wavelength of 500 nm or more can ensure transmission of ultraviolet light, visible light, and infrared light.
  • the detector has the same component composition as that of Embodiment 1 or the detector and the fluorescent chromatography detector commonly used in the prior art, except that the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • a fluorescent immunochromatography detecting device comprises a test strip and a detecting device
  • the test strip comprises a multilayer porous membrane and a supporting sheet
  • the multilayer porous membrane is a sample mat, a bonding mat, a nitrocellulose membrane and an absorbent paper, respectively a fluorescent labeling reagent is fixed on the bonding pad
  • the nitrocellulose membrane is provided with a detection line and a control line
  • the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet, thereby obtaining a test strip.
  • the support sheet is an ethylene glycol modified polyethylene terephthalate material.
  • the ethylene glycol-modified polyethylene terephthalate material has a light transmittance of 10% or less at a wavelength of less than 400 nm, and a light transmittance of 90% or more at a wavelength of 500 nm or more, which can ensure ultraviolet light, visible light, and Transmission of infrared rays.
  • the detector has the same component composition as that of Embodiment 1 or the detector and the fluorescent chromatography detector commonly used in the prior art, except that the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • a fluorescent immunochromatography detecting device comprises a test strip comprising a multi-layer porous membrane and a support sheet, and a multi-layer porous membrane respectively being a sample mat, a binding mat, a nitrocellulose membrane and a water absorbing material Paper, a fluorescent labeling reagent is fixed on the bonding pad, the nitrocellulose membrane is provided with a detection line and a control line, and the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet to obtain a test paper. article.
  • the support sheet is a cellulose acetate butyrate material.
  • the cellulose acetate butyrate material has a light transmittance of 10% or less at a wavelength of less than 400 nm, and a light transmittance of 90% or more at a wavelength of 500 nm or more, and can ensure transmission of ultraviolet light, visible light, and infrared light.
  • the detector has the same component composition as that of Embodiment 1 or the detector and the fluorescent chromatography detector commonly used in the prior art, except that the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • a fluorescent immunochromatography detecting device comprises a test strip and a detecting device
  • the test strip comprises a multilayer porous membrane and a supporting sheet
  • the multilayer porous membrane is a sample mat, a bonding mat, a nitrocellulose membrane and an absorbent paper, respectively a fluorescent labeling reagent is fixed on the bonding pad
  • the nitrocellulose membrane is provided with a detection line and a control line
  • the sample pad, the bonding pad, the nitrocellulose membrane and the absorbent paper are sequentially attached to each other on the support sheet, thereby obtaining a test strip.
  • the support sheet is a polyacrylate material.
  • the polyacrylate material has a light transmittance of 10% or less at a wavelength of less than 450 nm, and a material having a light transmittance of 90% or more at a wavelength of 500 nm or more, and can ensure transmission of ultraviolet light, visible light, and infrared light.
  • the detector has the same component composition as that of Embodiment 1 or the detector and the fluorescent chromatography detector commonly used in the prior art, except that the excitation light source and the photodetector are respectively located on both sides of the detection area.
  • the immunochromatographic detection device described in Example 1 was used for immunofluorescence detection of troponin, and the steps were as follows:
  • test strip and whole blood/serum/plasma specimens were returned to room temperature (20-30 ° C) before use, and the reaction buffer was taken out and allowed to stand for 20 minutes to return to room temperature.
  • test strip Place the test strip on a clean, flat surface, use a pipette to draw fingertip blood or venous blood, and then drop 1 drop of whole blood/serum/plasma (about 25 ml) onto the sample pad.
  • results of the detection using the immunochromatographic assay device of the present invention were compared with hospital correlation data.
  • the results are shown in Table 1 and Figure 4.
  • Figure 4 the detection using the apparatus of the present invention is illustrated.
  • the results have a good correlation with the hospital test results, and the accuracy of the apparatus of the present invention is relatively high.
  • Table 1 shows the results of detecting troponin using the immunochromatographic assay device of the present invention.

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Abstract

一种免疫层析检测装置，包括试纸条和检测装置，该试纸条包括多层多孔膜和支撑片，支撑片为在小于450nm波长具有10％以下透光率、在500nm波长以上具有90％以上透光率的材料；检测器包括激发光源、放置试纸条的检测区和光检测器，激发光源和光检测器分别位于检测区的两侧。该检测装置采用透射法，光检测器能够捕捉多层多孔膜表面和内部的荧光信号，使检测结果更加准确。

Description

一种免疫层析检测装置 技术领域
本发明涉及免疫分析技术领域，具体地，涉及一种免疫层析检测装置。
背景技术
免疫层析法(immunochromatography)是近几年来国外兴起的一种快速诊断技术，其原理是将特异的抗体先固定于硝酸纤维素膜的某一区带，当该干燥的硝酸纤维素一端浸入样品(尿液或血清)后，由于毛细管作用，样品将沿着该膜向前移动，当移动至固定有抗体的区域时，样品中相应的抗原即与该抗体发生特异性结合，若用免疫胶体金或免疫酶染色可使该区域显示一定的颜色，来提供对样品的定性或半定量检测数据，从而实现特异性的免疫诊断。与常规诊断方法相比，该技术分析速度快，全部检测过程仅需5～30分钟。操作简便，不需要其它任何仪器，无需专业人员，为实时实地检测提供了条件。检测样品种类多，可用于医疗、卫生、食品安全、环境检测中的血液、唾液、食物、水质、土壤等制备的微量溶液。
免疫层析试纸条是结合免疫标记物与对应抗原(抗体)的反应特异性和层析技术而制成的。免疫层析测试条通常由加样区、反应区和吸水区三部分组成。加样区含有免疫标记物颗粒，通常由玻璃纤维将免疫胶体金颗粒吸附在该区；反应区则喷涂两条反应线，一条为检测线，一条为质控线。检测线用以检测此处(抗原)物质与免疫标记物颗粒的包被抗原(抗体或抗体携带的抗原)的反应性；质控线则用以检测免疫标记物颗粒上的包被蛋白质的活性及程度，免疫标记物由加样区释放后，经反应区，部分继续到吸附区，完成层析，从而达到快速检测的目的。
最新的免疫层析法采用化学交联或生物分子间特异性作用将特异性抗体偶联到荧光分子或荧光颗粒的表面，分别得到荧光染料修饰的特异性抗体和荧光染料修饰的质控分子。当荧光染料表面存在活性基团时，可将其直接与特异性抗体反应，不需用化学交联剂；反之，则需通过化学交联剂将荧光染料与特异性抗体或质控分子相偶联。其中，化学交联剂包括1-乙基-3-(3-二甲基胺丙基)碳化二亚胺(EDC)、N-羟基琥珀酰亚胺(NHS)及戊二醛等。
为了提高信号与背景的区分度，荧光染料的荧光波长范围应为550～800nm。因为在紫外照射下，层析膜、底板和扣卡的自身荧光强度在550nm以下会远强于550nm以上，从而会对低浓度蛋白的检测产生一定的影响，故优选发射波长大于550nm的荧光染料；此外，层析膜、底板和扣卡一般在近红外区域(600～800nm)荧光强度极弱，因此，优选波长范围为550～800nm的荧光染料以进一步提高灵敏度。荧光染料包括有机荧光染料和稀土元素荧光染料，例如AlexaFluro系列的荧光素。
目前采用的免疫荧光层析装置采用反射法检测多孔膜上的荧光信号，荧光检测仪捕捉到的是多孔膜表面荧光染料修饰的特异性抗体，而检测不到多孔膜内部的荧光信号，导致检测灵敏度下降。
发明内容
本发明为了克服现有技术的上述不足，提供一种免疫层析检测装置，该装置创造性地采用透射法，光检测器能够捕捉多层多孔膜表面和内部的荧光信号，使检测结果更准确。
为了实现上述目的，本发明是通过以下技术方案予以实现的：
一种免疫层析检测装置，由试纸条和检测器组成，所述试纸条包括多层多孔膜和支撑片，支撑片为在小于450nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料；检测器腔体内部包括激发光源、放置试纸条的检测区和光检测器，所述激发光源和光检测器分别位于检测区的两侧。
本发明检测器的组成部件与传统的免疫层析检测器的部件组成完全相同，唯一不同之处在于，本发明检测器中的激发光源和光检测器分别位于检测区的两侧；而传统的检测器中的激发光源和光检测器位于检测区的同一侧。这种区别在于，传统的检测器是利用反射法检测多层多孔膜上的荧光信号，而本发明是通过透射法检测多层多孔膜上的荧光信号。为了配合透射法的实现，本发明将传统试纸条中的支撑片的材料进行了改进，使用了在小于450nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料。这种材料可以允许绝大多数的可见光透过。这样激发光源能穿透多层多孔膜的表面及内部，光检测器能够捕捉多层多孔膜表面和内部的荧光信号，使检测结果更准确。
根据支撑片对于不同波长光源的透过要求，支撑片可以是无色或者有色的， 塑料材质的支持物可掺入染料和颜料以吸引一定波长的光子，来满足对于不同波长光源的透过要求。优选地，所述支撑片为聚酯、亚克力、聚碳酸酯、聚丙烯酸酯、乙二醇改性的聚对苯二甲酸乙二醇酯或乙酸丁酸纤维素材料。
更优选地，所述支撑片为聚碳酸酯材料。聚碳酸酯是光学树脂材料中折光指数较高的品种，其透光率与无机玻璃相当，折射率达到1.59，密度为无机玻璃的一半，能自由成型为任意形状，具有良好的生物相容性。
与现有技术相比，本发明具有如下有益效果：
传统的免疫荧光层析方法为将一定量的样本滴加到样品垫上，通过层板作用向前移动，溶解在结合垫上的标记试剂与样本反应，当结合物移动至抗原检测带时，待测物与结合物发生特异性的反应被截留，结合物在检测线上富集，结合物含量与样品中的待测物含量成正比。在激发光源下产生的荧光强度与试纸条上的结合物含量成正比，当光源照射到试纸条的检测区和质控区时，激发附着的荧光物质，发射光收集并转化为电信号，电信号的强弱和荧光分子数量相关，检测仪计算样品中待测物的含量。透射法比传统的反射方法的优点在于灵敏度高。因为传统的反射方法只是采集膜片表面的抗体信号，而透射法连膜内部的抗体反应信号都能采集，所以信号会更高。
附图说明
图1为检测器的立体示意图。
图2为检测器腔体内部结构示意图。
图3为检测卡槽的结果示意图。
图4为与医院检查数据相关性比较结果。
具体实施方式
下面结合说明书附图和具体实施例对本发明作出进一步地详细阐述，所述实施例只用于解释本发明，并非用于限定本发明的范围。下述实施例中所使用的试验方法如无特殊说明，均为常规方法；所使用的材料、试剂等，如无特殊说明，为可从商业途径得到的试剂和材料。
实施例1
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水 纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得到试纸条。本实施例中支撑片为聚碳酸酯材料。聚碳酸酯材料在小于400nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器的立体示意图见图1，检测器腔体内部结构示意图见图2。检测器包括壳体和至于壳体内的激发光源、放置试纸条的检测卡槽、光检测器、微控制器，所述激发光源和光检测器分别位于检测卡槽的两侧，激发光源和光检测器通过检测电路与微控制器连接。检测器的壳体表面设有显示屏和一个卡槽开口，通过开口可以把检测卡槽从检测器壳体内抽出来，检测卡槽的抽出方式可以是抽拉式或者弹出式，检测卡槽为中间开口的回字形卡槽(见图3)，把试纸条放到检测卡槽上，试纸条的两头通过旋转压板或其他装置固定后，试纸条上的检测线和控制线正好落在口字形卡槽的中间开口处，这样激发光源通过试纸条的透射光可以没有任何阻挡被光检测器检测到。
本实施例所述荧光免疫层析检测装置的使用方法为，将一定量的待测样本滴加到样品垫上，通过层板作用向前移动，溶解在结合垫上的标记试剂与样本反应，当结合物移动至抗原检测线时，待测物与结合物发生特异性的反应被截留，结合物在检测线上富集，结合物含量与样品中的待测物含量成正比。在激发光源下产生的荧光强度与试纸条上的结合物含量成正比，微控制器控制激发光源照射到试纸条的检测线和控制线时，激发附着的荧光物质，光检测器收集通过试纸条上的透射光，并转化为电信号，电信号的强弱和荧光分子数量相关，电信号传送给微控制器，微控制器接收电流信号，进行转换处理分析，计算待测样品中待测物的含量，微控制器将处理分析的结果通过检测器壳体表面的显示屏显示出来。
实施例2
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得 到试纸条。本实施例中支撑片为亚克力材料。亚克力材料400nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器同实施例1或者检测器的组件组成和现有技术中常用的荧光层析检测器相同，唯一不同的是激发光源和光检测器分别位于检测区的两侧。
实施例3
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得到试纸条。本实施例中支撑片为聚酯材料。聚酯材料小于400nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器同实施例1或者检测器的组件组成和现有技术中常用的荧光层析检测器相同，唯一不同的是激发光源和光检测器分别位于检测区的两侧。
实施例4
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得到试纸条。本实施例中支撑片为乙二醇改性的聚对苯二甲酸乙二醇酯材料。乙二醇改性的聚对苯二甲酸乙二醇酯材料在小于400nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器同实施例1或者检测器的组件组成和现有技术中常用的荧光层析检测器相同，唯一不同的是激发光源和光检测器分别位于检测区的两侧。
实施例5
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水 纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得到试纸条。本实施例中支撑片为乙酸丁酸纤维素材料。乙酸丁酸纤维素材料在小于400nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器同实施例1或者检测器的组件组成和现有技术中常用的荧光层析检测器相同，唯一不同的是激发光源和光检测器分别位于检测区的两侧。
实施例6
一种荧光免疫层析检测装置，包括试纸条和检测装置，所述试纸条包括多层多孔膜和支撑片，多层多孔膜分别为样品垫、结合垫、硝酸纤维素膜和吸水纸，结合垫上固定有荧光标记试剂，硝酸纤维素膜设有检测线和控制线，样品垫、结合垫、硝酸纤维素膜和吸水纸顺次相互搭接地粘贴在支撑片上，从而得到试纸条。本实施例中支撑片为聚丙烯酸酯材料。聚丙烯酸酯材料在小于450nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料，能保证紫外光，可见光和红外线的透过。
检测器同实施例1或者检测器的组件组成和现有技术中常用的荧光层析检测器相同，唯一不同的是激发光源和光检测器分别位于检测区的两侧。
应用例1
使用实施例1所述的免疫层析检测装置用于肌钙蛋白的免疫荧光检测，步骤如下：
1、使用前将检测试纸条和全血/血清/血浆标本恢复至室温(20～30℃)，取出反应缓冲液并放置20分钟使其恢复至室温。
2、将试纸条置于干净平坦的台面上，用吸管吸取指尖血或静脉血后滴入1滴全血/血清/血浆(约25ml)于样品垫上。
3、在15分钟左右将试纸条放在检测器的检测卡槽中，检测器读取测试结果。
4、根据检测器数据进行结果判定。
将使用本发明所述免疫层析检测装置的检测结果与医院的相关性数据进行对比，结果见表1和图4。由图4可以看出，说明使用本发明所述仪器检测的 结果与医院检测结果有良好的相关性，本发明所述仪器的准确率比较高。
表1为使用本发明所述的免疫层析检测装置检测肌钙蛋白的结果
  T C T/C 医院检测值
样本1 8658 27225 0.3180165 1.34
样本2 19059 22275 0.8556229 1.69
样本3 35032 26586 1.317686 3.05
样本4 33687 18056 1.8656956 4.32
样本5 36309 26271 1.3820943 4.21
样本6 2770 22418 0.1235614 0.1
样本7 1982 24053 0.0824014 0.05
样本8 9949 35512 0.2801588 0.22
样本9 4328 27659 0.1564771 0.1
样本10 1236 18791 0.0657762 0.04
样本11 5416 42398 0.1277419 0.12
样本12 10882 52509 0.2072407 0.29
样本13 4453 30794 0.1446061 0.1
样本14 3704 21401 0.173076 0.05
样本15 10276 21266 0.4832126 0.2
样本16 5359 35452 0.1511621 0.15
样本17 3543 43317 0.0817924 0.05
样本18 3411 51402 0.0663593 0.05
样本19 6788 29765 0.2280531 0.27
样本20 3699 39170 0.0944345 0.07
样本21 6435 76371 0.0842597 0.07
样本22 4547 31546 0.1441387 0.09
样本23 4994 32618 0.1531056 0.18
  T C T/C 医院检测值
样本24 3658 22450 0.1629399 0.05
样本25 24603 48801 0.5041495 2.29
样本26 4709 41490 0.1134972 0.14
样本27 2160 15676 0.1377903 0.05
样本28 11802 16819 0.7017064 1.3
样本29 3874 29252 0.1324354 0.14
样本30 2559 24449 0.1046669 0.07
样本31 16233 21443 0.7570303 0.85
样本32 4453 33311 0.1336796 0.14
样本33 4727 22316 0.2118211 0.14
样本34 2814 34037 0.0826747 0.07
样本35 2567 16482 0.1557457 0.11
样本36 5877 15121 0.3886648 0.53
样本37 4694 16515 0.2842265 0.59
样本38 6953 23254 0.2990023 0.36
样本39 15043 23419 0.6423417 1.25
样本40 46214 35507 1.3015462 4.62
样本41 4375 26463 0.1653252 0.25
样本42 4588 35139 0.1305672 0.1
样本43 3765 35036 0.1074609 0.1
样本44 7929 24685 0.3212072 1.33
样本45 13161 61423 0.2142683 1.29
样本46 4506 53640 0.0840045 0.06

Claims (3)

  1. 一种免疫层析检测装置，由试纸条和检测器组成，所述试纸条包括多层多孔膜和支撑片；检测器腔体内部包括激发光源、放置试纸条的检测区和光检测器，其特征在于，所述试纸条的支撑片为在小于450nm波长具有10％以下透光率，在500nm波长以上具有90％以上的透光率的材料；所述激发光源和光检测器分别位于检测区的两侧。
  2. 根据权利要求1所述的装置，其特征在于，所述支撑片为聚酯、亚克力、聚碳酸酯、聚丙烯酸酯、乙二醇改性的聚对苯二甲酸乙二醇酯或乙酸丁酸纤维素材料。
  3. 根据权利要求1所述的装置，其特征在于，所述支撑片为聚碳酸酯材料。
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