Immunodetection reagent card based on photonic crystal fluorescence enhancement

Abstract

The utility model discloses an immunodetection reagent card based on photonic crystal fluorescence enhancement. The immunodetection reagent card comprises a shell, a biochip and a gland, the shell is enclosed by an upper shell and a lower shell, the upper shell is provided with a detection window, and at least two photonic crystals of the biochip are exposed out of the shell through the detection window; the gland is at least provided with a reaction sample adding hole and a quality control sample adding hole corresponding to the positions of the two photonic crystals respectively; the gland can be attached to the biochip, so that the inner wall of the reaction sample adding hole and the inner wall of the quality control sample adding hole are enclosed with the surface of the biochip to form a reagent reaction bin and a reagent quality control bin respectively. The method has the advantages that 1) high-sensitivity detection of ultralow-concentration substances can be realized, and the detection accuracy is improved; 2) a sample pad, a gold label pad, a chromatography membrane and other components are omitted, the production process is more convenient, and the production cost is reduced; and 3) mutual crosstalk of samples is effectively avoided, the efficiency of immunoreaction is ensured, and the accuracy of a detection result is improved.

Description

TECHNICAL FIELD

[0001] The utility model relates to a kind of in-vitro immune detection technology, especially a kind of immune detection reagent card. BACKGROUND

[0002] Traditional immunofluorescence detection technology has limited sensitivity when detecting ultra-low concentration substances, making it difficult to meet the needs of some high-precision detection applications. Existing immunochromatographic reagent cards (CN108872412A, CN118050505B) usually have complex structures and generally use a multi-layer membrane structure (sample pad, conjugate pad, chromatographic membrane, and absorbent pad). However, they have the following technical defects:

[0003] Structural redundancy: The multi-layer membrane assembly requires precise assembly, resulting in low production yield (usually <85%) and high material cost (over 60%);

[0004] Sensitivity bottleneck: Relies on colloidal gold / flourescent microsphere labeling, with detection limit limited to ng / mL level, unable to meet the needs of ultra-low concentration index detection such as ctDNA;

[0005] Signal interference: Non-specific adsorption of sample matrix during chromatography is severe, resulting in high background noise (signal-to-noise ratio usually <10:1).

[0006] Existing immunochromatographic reagent cards not only increase the complexity of production process, but also affect the accuracy and sensitivity of detection to some extent. Therefore, it is of great practical significance to develop a test reagent card that can achieve ultra-low concentration sensitivity detection and has a simple structure. INVENTION CONTENTS

[0007] To simplify the immune test reagent card while achieving ultra-low concentration sensitivity detection, the utility model provides an immune test reagent card based on photonic crystal fluorescence enhancement.

[0008] The utility model solves its technical problems by adopting the following technical scheme: an immune test reagent card based on photonic crystal fluorescence enhancement, including a shell, a biochip fixed in the shell; further including a gland; the shell is enclosed by an upper shell and a lower shell, the upper shell is provided with a detection window, at least two photonic crystals of the biochip are exposed outside the shell through the detection window; the gland is provided with a reaction sample hole and a quality control sample hole corresponding to the positions of the two photonic crystals, respectively; and the gland can be attached to the biochip, so that the inner walls of the reaction sample hole and the quality control sample hole and the surface of the biochip form a reagent reaction chamber and a reagent quality control chamber, respectively.

[0009] It is easily understood that the specific way of enclosing the upper shell and the lower shell to form the shell can be selected by common methods, for example, the following scheme can be adopted: the upper shell is provided with a plurality of upper conical holes, and the corresponding position of the lower shell is provided with a cylindrical pin matched with the upper conical hole; when the upper shell and the lower shell are closed, the cylindrical pin is inserted into the upper conical hole, and a pin positioning structure is formed by interference fit to realize the enclosure of the upper shell and the lower shell. In addition, the upper clamping edge and the lower clamping edge matched with each other can also be arranged on the opposite edges of the upper shell and the lower shell respectively, and the upper clamping edge and the lower clamping edge are clamped with each other to realize the enclosure of the upper shell and the lower shell. The two enclosure structures can also be used at the same time to increase the firmness and tightness of the enclosure.

[0010] It is easily understood that the micro-gap lamination of the gland and the biochip can also adopt various common locking structures, for example, we provide a preferred scheme: the gland is provided with a locking buckle, and the upper shell or the lower shell is provided with a locking groove matched with the locking buckle; when the gland is laminated to the surface of the biochip, the locking buckle and the locking groove can be locked with each other to realize the micro-gap lamination of the gland and the biochip. The above-mentioned "micro-gap lamination" means that a capillary gap is left on the lamination surface of the gland and the biochip, which cooperates with the capillary stop groove below to form a step with a size mutation, constituting a capillary stop valve to prevent liquid flow (see Figure 11 ). For example, as shown in the specific embodiment, as a preferred scheme, the locking buckle can be connected with the gland body through an elastic arm, and the elastic arm can quickly realize the buckling and separation of the locking buckle and the locking groove.

[0011] As a further improvement of the utility model, the lamination surface outside the non-reaction area of the gland and the biochip is further provided with a capillary stop groove for preventing the sample reaction liquid from breaking through the micro-gap lamination surface and flowing to the non-reaction area, and the depth of the capillary stop groove is not less than 1 mm.

[0012] As a further improvement of the utility model, it further includes a waste liquid absorbing pad for absorbing the waste liquid generated in the detection process, and a water absorbing pad mounting chamber is formed in the inside of the shell for mounting the waste liquid absorbing pad.

[0013] As a further improvement of the utility model, a waste liquid guide groove for guiding the waste liquid is arranged between the detection window and the water absorbing pad mounting chamber, so that the waste liquid generated in the detection process can smoothly enter the water absorbing pad.

[0014] As a further improvement of the utility model, the surface of the upper shell is further provided with an identification groove for mounting a drying paper, and the drying paper can be used to wipe the liquid remaining on the surface of the photonic crystal.

[0015] As a further improvement of the utility model, the diameters of the reaction sample adding hole and the quality control sample adding hole are 2.5-3.5 times of the depths of the reagent reaction bin and the reagent quality control bin respectively.

[0016] As a further improvement of the utility model, the substrate of the biochip is PET material. The reagent card only adopts one layer of PET substrate, and the photonic crystal is directly printed on the substrate, compared with the traditional immunochromatography reagent card, the sample pad, the gold mark pad and the chromatography film and other components are reduced, so that the overall structure is simpler, the production process is more convenient, and the production cost is reduced. Specifically, the photonic crystal array can be composed of 3*3 micro points, and the size of each micro point is 400-600 um, and the micro point spacing is less than 700 um.

[0017] The utility model has the advantages that: 1) the utility model utilizes the enhancement function of photonic crystal to specific wavelength fluorescent signal, combines traditional immunofluorescence detection, can realize the high sensitivity detection of ultralow concentration material, improves the accuracy and reliability of detection;Black PET substrate reduces background noise, combined with the local field enhancement effect of photonic crystal, makes the detection limit reach 0.1fg / mL (improves 10 4 times compared with colloidal gold method). 2) cancel the sample pad, gold mark pad and chromatography film of traditional reagent card, adopt "single layer PET substrate + double reaction cavity" integrated design, reduce more than 80% production consumables;Replace nitrocellulose membrane with photonic crystal micro column array, avoid batch difference caused by uneven chromatography flow rate. 3) the detection cavity and quality control cavity of the reagent card are independent of each other, avoid sample mutual interference, ensure the efficiency of immunoreaction, further improve the accuracy of detection result. BRIEF DESCRIPTION OF DRAWINGS

[0018] Figure 1 It is the immunological detection reagent card structure diagram (explosion diagram) of the embodiment based on photonic crystal fluorescence enhancement.

[0019] Figure 2 It is the top shell top view of the embodiment.

[0020] Figure 3 It is the top shell bottom view of the embodiment.

[0021] Figure 4 It is the top shell section view of the embodiment.

[0022] Figure 5 It is the bottom shell top view of the embodiment.

[0023] Figure 6 It is the bottom shell side structure diagram of the embodiment.

[0024] Figure 7 It is the biochip structure diagram of the embodiment.

[0025] Figure 8 is a top view of the gland of the specific embodiment.

[0026] Figure 9 is a sectional view of the gland of the specific embodiment.

[0027] Figure 10 is a bottom view of the gland of the specific embodiment.

[0028] Figure 11 is a schematic view of the capillary stop valve structure formed by the surface of the gland and the biochip and the capillary stop groove of the specific embodiment.

[0029] In the figure, the marks are: 1-biochip, 101-substrate, 102-photonic crystal, 2-gland, 201-reaction sample hole, 202-quality control sample hole, 203-locking buckle, 204-capillary stop groove, 205-elastic arm, 3-upper shell, 301-detection window, 302-upper conical hole, 303-upper buckling edge, 304-waste liquid guide groove, 305-identification groove, 306-pressing edge, 4-lower shell, 401-cylindrical pin, 402-lower buckling edge, 403-locking groove, 404-chip groove, 5-waste liquid absorbent pad, 6-absorbent pad mounting chamber, 7-drying paper. Specific embodiment

[0030] The utility model is further explained in connection with the drawings and examples.

[0031] As Figures 1-10 shown, the utility model discloses an immunological detection reagent card based on photonic crystal fluorescence enhancement, including the casing, the biochip 1 (fix through chip groove 404 and pressing edge 306) and the gland 2 fixed in casing, the substrate 101 of biochip 1 is PET, the casing is enclosed by upper shell 3 and lower shell 4, the upper shell 3 is set with detection window 301, and two photonic crystals 102 of biochip 1 are exposed to the outside of casing through detection window 301, the gland 2 is set with reaction sample hole 201 and quality control sample hole 202 respectively corresponding with the position of two photonic crystals 102, and the gland 2 can be attached with biochip 1, so that the inner wall of reaction sample hole 201 and the inner wall of quality control sample hole 202 and the surface of biochip 1 form reagent reaction bin and reagent quality control bin respectively, the diameter of reaction sample hole 201 and quality control sample hole 202 is 3 times the depth of reagent reaction bin and reagent quality control bin respectively, and the surface of upper shell 3 is further provided with identification groove 305 for installing drying paper 7.

[0032] The upper shell 3 is provided with a plurality of upper conical holes 302, and the lower shell 4 is provided with a cylindrical pin 401 matched with the upper conical hole 302; when the upper shell 3 and the lower shell 4 are closed, the cylindrical pin 401 is inserted into the upper conical hole 302, a pin positioning structure is formed by interference fit, and the upper clamping edge 303 and the lower clamping edge 402 matched with each other are respectively arranged on the opposite edges of the upper shell 3 and the lower shell 4, so that the upper shell 3 and the lower shell 4 are clamped by the upper clamping edge 303 and the lower clamping edge 402 to realize the enclosure.

[0033] The grommet 2 is provided with a locking buckle 203 connected with the grommet body through an elastic arm 205, and the upper shell 3 or the lower shell 4 is provided with a locking groove 403 matched with the locking buckle 203; when the grommet 2 is attached to the surface of the biochip 1, the locking buckle 203 and the locking groove 403 can be locked with each other to realize the micro-gap attachment of the grommet 2 and the biochip 1. The capillary cutoff groove 204 for preventing the sample reaction liquid from breaking through the micro-gap attachment surface and flowing to the non-reaction area is further arranged outside the attachment surface of the grommet 2 and the biochip 1, and the depth of the capillary cutoff groove 204 is 2 mm.

[0034] The immunological detection reagent card of the embodiment further includes a waste liquid absorbing pad 5 for absorbing the waste liquid generated in the detection process, and a pad mounting chamber 6 for mounting the waste liquid absorbing pad 5 is formed in the inside of the shell. The waste liquid guide groove 304 for guiding the waste liquid is arranged between the detection window 301 and the pad mounting chamber 6.

[0035] The detection process is as follows:

[0036] (1) Sample processing: collect blood samples and perform filtration and dilution treatment.

[0037] (2) Sample loading: accurately drop the treated blood samples into the reagent reaction bin and the reagent quality control bin respectively by using a 20ul quantitative pipette.

[0038] (3) Incubation: place the reagent card into a 37° incubation chamber for 10 minutes.

[0039] Cleaning and drying: take out the reagent card from the incubation chamber, place it at an angle of 45 degrees, and drop the cleaning liquid into the reagent reaction bin by squeezing the cleaning bottle, then absorb the cleaning liquid with a water-absorbing paper after flushing.

[0040] (4) Signal reading: insert the cleaned reagent card into the instrument to read the fluorescence signal intensity, and calculate the concentration of the target substance in the sample according to the pre-established standard curve.

Claims

1. An immunodetection reagent card based on photonic crystal fluorescence enhancement, comprising a housing, a biochip (1) fixed in the housing; characterized in that: Further comprising a cover (2); the shell is enclosed by an upper shell (3) and a lower shell (4), the upper shell (3) is provided with a detection window (301), at least two photonic crystals (102) of the biochip (1) are exposed outside the shell through the detection window (301); the cover (2) is provided with at least a reaction sample hole (201) and a quality control sample hole (202) corresponding to the positions of the two photonic crystals (102) respectively; and the cover (2) can be attached to the biochip (1), so that the inner wall of the reaction sample hole (201) and the inner wall of the quality control sample hole (202) and the surface of the biochip (1) form a reagent reaction chamber and a reagent quality control chamber respectively.

2. The photonic crystal-based fluorescence-enhanced immunoassay test strip of claim 1, wherein: The specific structure of the upper shell (3) and the lower shell (4) enclosing the shell includes: the upper shell (3) is provided with a plurality of upper conical holes (302), and the corresponding position of the lower shell (4) is provided with a cylindrical pin (401) matched with the upper conical hole (302); when the upper shell (3) and the lower shell (4) are closed, the cylindrical pin (401) can be inserted into the upper conical hole (302), and a pin positioning structure is formed by interference fit to realize the enclosure of the upper shell (3) and the lower shell (4).

3. The photonic crystal-based fluorescence-enhanced immunoassay test strip of claim 1, wherein: The specific structure of the upper shell (3) and the lower shell (4) enclosing the shell includes: the upper shell (3) and the lower shell (4) are respectively provided with upper and lower buckling edges (303) and (402) matched with each other on the opposite edges, and the upper and lower buckling edges (303) and (402) are buckled to each other to realize the enclosure of the upper shell (3) and the lower shell (4).

4. The photonic crystal-based fluorescence-enhanced immunoassay test strip of claim 1, wherein: The specific structure of the cover (2) and the biochip (1) is that the cover (2) is provided with a locking buckle (203), and the upper shell (3) or the lower shell (4) is provided with a locking groove (403) matched with the locking buckle (203); when the cover (2) is attached to the surface of the biochip (1), the locking buckle (203) and the locking groove (403) can be locked to each other to realize the micro-gap attachment of the cover (2) and the biochip (1).

5. The photonic crystal-based fluorescence-enhanced immunoassay test strip of claim 4, wherein: A capillary cutoff groove (204) is further arranged outside the non-reaction area of the attachment surface of the cover (2) and the biochip (1) to prevent the reaction liquid from flowing to the non-reaction area through the micro-gap attachment surface, and the depth of the capillary cutoff groove (204) is not less than 1mm.

6. The immunodetection reagent card based on photonic crystal fluorescence enhancement according to any one of claims 1 to 5, characterized by: Further comprising a waste liquid absorbing pad (5) for absorbing waste liquid generated in the detection process, and a water absorbing pad mounting chamber (6) is formed in the inside of the shell for mounting the waste liquid absorbing pad (5).

7. The photonic crystal-based fluorescence-enhanced immunoassay test strip of claim 6, wherein: A waste liquid flow guide groove (304) is arranged between the detection window (301) and the water absorbing pad mounting chamber (6) for guiding the waste liquid.

8. The immunodetection reagent card based on photonic crystal fluorescence enhancement according to any one of claims 1 to 5, characterized by: An identification groove (305) is further arranged on the surface of the upper shell (3) for mounting a drying paper (7).

9. The immunodetection reagent card based on photonic crystal fluorescence enhancement according to any one of claims 1 to 5, characterized by: The diameters of the reaction sample hole (201) and the quality control sample hole (202) are 2.5-3.5 times the depth of the reagent reaction chamber and the reagent quality control chamber respectively.

10. The immunoassay test strip based on photonic crystal fluorescence enhancement according to any one of claims 1 to 5, characterized in that: The substrate (101) of the biochip (1) is PET material.

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