Fucose glycoprotein detection kit, preparation method thereof and fucose glycoprotein coloration method

Abstract

The application discloses a fucose glycoprotein detection kit, a preparation method thereof and a fucose glycoprotein color developing method. First, a liquid sample is dropped on the surface of the kit, and the sample and test paper are infiltrated through a sample inlet. Under the action of transverse flow force, the sample flows to an observation window along a hydrophilic test paper. The glycoprotein containing alpha 1,2-fucose in the sample is combined with agglutinin 1, and the rest of the sample continues to flow to the next layer of test paper. Then, the glycoprotein containing alpha 1,3-fucose is combined with agglutinin 2, and the rest of the sample flows to the third layer and is combined with agglutinin 3 containing alpha 1,6-fucose glycoprotein. All the combined fucose glycoproteins are combined with agglutinin biotin through their antibodies, and horseradish peroxidase is used for labeling color development, so that quantitative detection of the fucose glycoprotein in the sample is realized. The method has important significance for studying the changes of different connection fucose glycoproteins in cells and body fluids in diseases.

Description

Technical Field

[0001] This invention belongs to the field of biomolecular analysis reagent technology, specifically relating to a fucoglycoprotein colorimetric method and a reagent kit using this colorimetric method and its preparation method. Background Technology

[0002] Fucosylation of proteins is highly expressed in various tumors, and abnormal fucosylation indicates tumor development and progression. Literature review indicates that the types and abundance of core and branched fucose in bovine peptide protein (AFP) can differentiate the malignancy of hepatitis and liver cancer; prostate-specific antigen (PSA) fucosylation, a commonly used biomarker, can be used to differentiate the invasiveness of prostate cancer; numerous studies have shown that protein fucosylation predicts malignant metastasis and spread in gastric, pancreatic, and rectal cancers. Therefore, quantitative detection of protein fucosylation in body fluids is of great significance for tumor screening, diagnosis, and prognosis. Summary of the Invention

[0003] The purpose of this invention is to provide a fucoidan glycoprotein detection kit, its preparation method, and a fucoidan glycoprotein colorimetric method, so as to solve the problem of quantitatively detecting specific fucoidylated glycoproteins in the prior art.

[0004] One technical solution of the present invention is:

[0005] A method for preparing a fucoidan glycoprotein detection kit, comprising the following steps:

[0006] (1) Select hydrophilic materials containing cellulose as hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3, the lengths of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 increase sequentially, and set one end of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 as lectin binding region respectively;

[0007] (2) Immerse the lectin binding regions of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 in the oxidizing solution and react at room temperature for 1-2 hours, then remove them;

[0008] (3) The lectin-binding region of the hydrophilic material 1 is immersed in the first lectin solution, the lectin-binding region of the hydrophilic material 2 is immersed in the second lectin solution, and the lectin-binding region of the hydrophilic material 3 is immersed in the third lectin solution. All are reacted at room temperature for 6-12 h to obtain the first lectin-binding material, the second lectin-binding material and the third lectin-binding material, respectively.

[0009] (4) Take out the first lectin binding material, the second lectin binding material and the third lectin binding material, clean them, and vacuum dry the three lectin binding materials at room temperature.

[0010] (5) Align one end of the first lectin bonding material, the second lectin bonding material and the third lectin bonding material to form an aligned end, and bond the first lectin bonding material, the second lectin bonding material and the third lectin bonding material in order from top to bottom, with the lectin bonding areas all exposed. Then attach a support base plate to the bottom of the third lectin bonding material and attach the fixing material to the support base plate, at which time the fixing material is close to the lectin bonding area.

[0011] (6) Fix the bottom plate of the box to the bottom of the support base plate, and cover the box cover on the bottom plate to obtain the fucose detection kit. The box cover has a sample injection cell and an observation window. The sample injection cell is located above the alignment end, and the observation window is located above the lectin binding region.

[0012] Furthermore, in step (2), the oxidizing solution is 10-20 mM NaIO3 dissolved in 50-100 mM CH3COONa buffer solution, pH = 4-6.

[0013] Furthermore, in step (3), the first lectin in the first lectin solution is any one or more of UEA-I, PTL-II, and LAA; the second lectin in the second lectin solution is LTL; and the third lectin in the third lectin solution is any one or two of LCH and AAL.

[0014] Furthermore, in step (4), the cleaning specifically involves immersing the device in 20-30% acetonitrile for 2-4 times, followed by rinsing with deionized water for 3-5 times.

[0015] Another technical solution of the present invention is:

[0016] A method for preparing a fucoidan glycoprotein detection kit. The fucoidan glycoprotein detection kit includes: a box cover and a box base plate, the box cover and the box base plate forming a hollow receiving cavity. A sample injection cell and an observation window are provided on the box cover. A support base plate is fixed on the box base plate. A third lectin binding material, a second lectin binding material and a first lectin binding material are sequentially fixed on the support base plate from bottom to top. One end of the third lectin binding material, the second lectin binding material and the first lectin binding material are aligned to form an aligned end, and are located below the sample injection cell. The other ends of the third lectin binding material, the second lectin binding material and the first lectin binding material decrease in size in a stepped shape, so that the lectin binding area is located below the observation window.

[0017] Furthermore, the thickness of the third lectin-binding material, the second lectin-binding material, and the first lectin-binding material is 0.5-1.0 mm, and the length of the third lectin-binding material, the second lectin-binding material, and the first lectin-binding material decreases by 5-10 mm in sequence.

[0018] Another technical solution of the present invention is:

[0019] A colorimetric method for fucoidan glycoprotein detection in a fucoidan assay kit includes:

[0020] (1) The sample was thoroughly mixed with fucoidan antibody, lectin biotin and HRP (horseradish peroxidase) at room temperature to obtain a mixed sample;

[0021] (2) The mixed sample is dropped into the sample injection cell. The mixed sample comes into contact with the hydrophilic material No. 1 and flows laterally to the color development area under the observation window. It reacts with the lectin binding area of ​​the first lectin binding material. When the mixed sample contains α1,2-fucoprotein, α1,2-fucoprotein binds to the first lectin, and the antibody-lectin-biotin-HRP in the mixed sample develops color, forming the first band. When the mixed sample does not contain α1,2-fucoprotein, the antibody-lectin-biotin-HRP in the mixed sample does not develop color.

[0022] (3) The mixed sample flows downward, so that the mixed sample comes into contact with the hydrophilic material No. 2, and reaches the color development area under the observation window through lateral flow, and reacts with the lectin binding area of ​​the second lectin binding material. When the mixed sample contains α1,3-fucoprotein, α1,3-fucoprotein binds to the second lectin, and the antibody-lectin-biotin-HRP in the mixed sample develops color, forming a second band. When the mixed sample does not contain α1,3-fucoprotein, the antibody-lectin-biotin-HRP in the mixed sample does not develop color.

[0023] (4) The mixed sample continues to flow downwards, so that the mixed sample comes into contact with the hydrophilic material No. 3, and reaches the color development area under the observation window through lateral flow, and reacts with the lectin binding area of ​​the third lectin binding material. When the mixed sample contains α1,6 fucoprotein, α1,6 fucoprotein binds to the third lectin, and the antibody-lectin biotin-HRP in the mixed sample develops color, forming a third band. When the mixed sample does not contain α1,6 fucoprotein, the antibody-lectin biotin-HRP in the mixed sample does not develop color.

[0024] (5) Let stand for 10-15 minutes to form a stable three-band result. The type and abundance of fucose glycoprotein contained in the sample are qualitatively and quantitatively determined by the color intensity.

[0025] Furthermore, after step (1) and before step (2), the method further includes: adding a reagent to the mixed sample and incubating the mixed sample and the reagent at room temperature for 5-15 minutes, wherein the reagent is prepared by dissolving DAB (diaminobenzidine) in 1×PBS buffer, wherein the weight percentage of DAB is 0.03-0.05%, and then adding CoCl2 at a weight percentage of 1.0-1.2%.

[0026] This invention provides a fucoidan glycoprotein detection kit, its preparation method, and a fucoidan glycoprotein colorimetric method. This invention can collect bodily fluids, including saliva, urine, and blood, and use fucoidan and its antibody, along with horseradish peroxidase colorimetric detection, to qualitatively and quantitatively detect fucosylation in bodily fluids, thereby achieving early screening and providing auxiliary diagnostic information for prognosis. Its advantages include:

[0027] (1) Using this kit, different linked fucoglycoproteins can be detected through saliva samples, making it convenient and quick to quantify the content of fucoglycoproteins;

[0028] (2) Quantitative testing of different linked fucoglycoproteins in cells;

[0029] (3) By extracting proteins from tissue samples, this kit is used to test and quantify different linked fucoglycoproteins, and to rapidly quantify the content and composition of fucoglycoproteins.

[0030] (4) Liquid samples containing fucoglycoprotein can be used directly for quantitative detection without pretreatment. Attached Figure Description

[0031] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0032] Figure 1 This is a schematic diagram of the fucoidan glycoprotein color development method and the kit preparation process using this color development method in this invention;

[0033] Figure 2 This is a schematic diagram of the fucoidan glycoprotein colorimetric method and the process of testing liquid samples using the kit based on this colorimetric method in this invention;

[0034] Figure 3This is a schematic diagram illustrating the principle of the HRP colorimetric method in the fucose glycoprotein colorimetric method of this invention;

[0035] Figure 4 This is a schematic diagram showing the relationship between the intensity of the three bands in the HRP colorimetric method of the present invention and the connection and content of fucose glycoprotein, wherein (a) is a sample without fucose; (b) is a sample containing α1,3 and α1,6 fucose; and (c) is a sample containing α1,2 / α1,3 / α1,6 fucose.

[0036] Figure 5 This is a schematic diagram illustrating the chemical binding process of fucoglycoprotein lectin on the surface of the test paper, the binding of fucoglycoprotein with lectin, and the binding process of fucoglycoprotein with antibody in this invention.

[0037] Figure 6 This is a schematic diagram illustrating the chemical binding process on the surface of the sialic acid glycoprotein lectin test paper, the binding process of fucoglycoprotein to lectin, and the binding process of fucoglycoprotein to antibody in this invention.

[0038] Figure 7 This is a schematic diagram illustrating the chemical binding process of T / Tn antigen glycoprotein lectin on the surface of the test strip, the binding of fucose glycoprotein with lectin, and the binding process of fucose glycoprotein with antibody in this invention. Detailed Implementation

[0039] This invention develops a colorimetric method for fucoidan glycoprotein, a reagent kit using this colorimetric method, and a manufacturing process.

[0040] Please see Figure 1 , Figure 1 This is a schematic diagram illustrating the fucoidan glycoprotein colorimetric method of the present invention and the preparation process of the reagent kit using this colorimetric method. Figure 1As shown, the solid-phase carrier consists of three layers of hydrophilic test paper (top layer hydrophilic material 1, middle layer hydrophilic material 2, and bottom layer hydrophilic material 3). The leading edge region of each hydrophilic material layer is used for covalent binding of lectins. After the three lectins are respectively bound to the three layers of hydrophilic test paper (hydrophilic material 1 with lectin 1, hydrophilic material 2 with lectin 2, and hydrophilic material 3 with lectin 2),... The reagents are prepared by bonding the three layers of lectin (number 3) to form a test strip layer (one end of the three layers aligned, the other end forming a stepped shape, with the first layer being the shortest and the third layer the longest). The bottom of the test strip layer is then attached to the support base plate and fixed to the bottom carrier (box bottom plate). Finally, the fixing material is attached to the support base plate, at which point the fixing material is close to the lectin end (lectin binding area) of the three layers of lectin. The final test kit is then packaged and installed. In the above structure, the thickness of each layer of hydrophilic test paper is 0.5-1.0 mm; the length of each layer of test paper increases by 5-10 mm. The test kit has a sample injection cell and an observation window at the top. The sample injection cell is located above the alignment end, and the observation window is located above the lectin end. The center of the sample injection cell is 2-3 cm away from the edge of the observation window.

[0041] The spacing between L1 / L2 / L3 is 0.5-1.0 cm.

[0042] In the above structure, the hydrophilic test paper uses a hydrophilic material containing cellulose, and the lectin-binding region of the hydrophilic test paper is immersed in an oxidizing agent (10-20 mM NaIO3 dissolved in 50-100 mM CH3COONa buffer solution, pH=4-6) for 1-2 hours at room temperature. The lectin-binding region of the hydrophilic test paper is then immersed in different lectin solutions: hydrophilic material 1 binds lectin 1 (UEA-I, PTL-II, LAA or one); hydrophilic material 2 binds lectin 2 (LTL); and hydrophilic material 3 binds lectin 3 (LCH, AAL or one), for 6-12 hours at room temperature. The lectin solution is 1×PBS buffer (pH 7.2-7.4). First, second, and third lectin-binding materials are obtained respectively.

[0043] The three lectin-binding materials were removed and rinsed 2-4 times with 20-30% acetonitrile, followed by 3-5 rinses with deionized water. The three lectin-binding materials were then vacuum-dried at room temperature. Please refer to the following: Figure 1 ,like Figure 1As shown, a layer of methylcellulose adhesive is applied to the upper and lower surfaces of the left side of the lectin-binding region of the second lectin-binding hydrophilic material. The three layers of lectin-binding material are then bonded together under pressure. The resulting three layers of lectin-binding material are then fixed to the next supporting base plate, and the fixing material is then fixed to the supporting base plate, thus securing the three layers of lectin-binding material in the test box.

[0044] Please refer to the procedure for testing liquid samples using the above kit. Figure 2 , Figure 2 This is a schematic diagram illustrating the fucoidan glycoprotein colorimetric method of the present invention and the process of testing liquid samples using a kit employing this method. Figure 2 As shown:

[0045] (a) Mix the liquid sample, fucoidan-specific antibody and lectin biotin (horseradish peroxidase) to obtain a mixed sample, and drop the mixed sample onto the aligned end of the hydrophilic material;

[0046] (b) The mixed sample flows laterally through a hydrophilic solid carrier, and the fucoidan binds to the three layers of hydrophilic test paper respectively;

[0047] (c) The lectin binds to the antibody and biotin, and binds to different linked fucoglycoproteins on the three-layer lectin test paper;

[0048] (d) The bound fucoprotein binds to biotin-horseradish peroxidase (HRP) and forms a color band that is related to the fucoprotein content.

[0049] The above-mentioned fucoglycoprotein binding and color development process is as follows: At room temperature, the sample is thoroughly mixed with fucoglycoprotein antibody, lectin biotin and HRP. The binding order is as follows: the test strip binds to the lectin, the lectin binds to the fucoglycoprotein in the sample, the fucoglycoprotein binds to the antibody, the antibody binds to the lectin biotin, and the lectin biotin binds to HRP.

[0050] Specifically as follows:

[0051] The mixed sample comes into contact with the carrier (hydrophilic material) and flows laterally from left to right, binding to the lectins. The carrier must be hydrophilic and possess a capillary structure to accelerate the rapid and sufficient flow of the liquid sample. During this lateral flow, α1,2-fucoproteins first bind to lectin 1, and the antibody-lectin-biotin-HRP attached to them develops color, forming the first band; if α1,2-fucoproteins are absent, no color develops. The remaining sample continues to flow to the next carrier layer, where α1,3-fucoproteins bind to lectin 2, and the antibody-lectin-biotin-HRP attached to them develops color, forming the second band; if α1,3-fucoproteins are absent, no color develops. The remaining sample continues to flow to the next carrier layer, where α1,6-fucoproteins bind to lectin 3, and the antibody-lectin-biotin-HRP attached to them develops color, forming the third band; if α1,6-fucoproteins are absent, no color develops. The reaction takes 10-15 minutes to form stable three-band results. The type and abundance of fucose glycoproteins in the sample are qualitatively and quantitatively determined by the color intensity. Please refer to [link to relevant documentation]. Figure 4 , Figure 4 This diagram illustrates the relationship between the intensity of the three HRP bands and the connectivity and content of fucoglycoprotein in the fucoglycoprotein colorimetric method of this invention. (a) represents a sample without fucose; (b) represents a sample containing α1,3 and α1,6 fucose; and (c) represents a sample containing α1,2 / α1,3 / α1,6 fucose. Figure 4 As shown, depending on the content of the fucoidan glycoprotein, the kit will show 1-3 bands that are positively correlated with the content.

[0052] Please refer to the above steps for the fucoglycoprotein horseradish peroxidase HRP colorimetric assay. Figure 3 , Figure 3 This is a schematic diagram illustrating the principle of the HRP colorimetric method in the fucose glycoprotein colorimetric method of this invention. Figure 3 As shown, (a) fucoprotein bound to the carrier is immobilized on the carrier by lectin. Fucoprotein binds to the antibody, and then to lectin-biotin and horseradish peroxidase (HRP); (b) DAB (diaminobenzidine) and hydrogen peroxide are added to the carrier. For example, 0.03-0.05% by weight of diaminobenzidine (DAB) is dissolved in 1×PBS buffer, and 1.0-1.2% by weight of CoCl2 (cobalt(II) chloride) is added. The mixed sample and reagents are incubated at room temperature for 5-15 minutes. HRP catalyzes the transfer of two electrons from lectin-biotin to hydrogen peroxide, which, together with water, produces an oxidized substrate, thereby developing color and forming a color band that is linearly related to the concentration.

[0053] To make the above-mentioned objects, features, and advantages of the present invention more apparent and understandable, the technical solutions of the present invention will be further described below in conjunction with the accompanying drawings and embodiments. However, the present invention is not limited to the listed embodiments, but should also include any other known modifications within the scope of the claims of the present invention.

[0054] First, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places throughout this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.

[0055] Secondly, this invention is described in detail using structural diagrams, etc. When detailing the embodiments of this invention, for ease of explanation, the diagrams may be partially enlarged, deviating from the general scale. Furthermore, the diagrams are merely examples and should not limit the scope of protection of this invention. In addition, actual manufacturing should include three-dimensional space with length, width, and depth.

[0056] Example 1

[0057] Fucose glycoprotein enrichment and quantification

[0058] Please see Figure 5 , Figure 5 This diagram illustrates the chemical binding process of fucoglycoprotein lectin on the surface of the test paper, the binding of fucoglycoprotein to lectin, and the binding of fucoglycoprotein to antibody in this invention. Figure 5 As shown, the surface of the hydrophilic test strip has groups (aldehyde or ketone groups) that can react with amino groups, allowing lectins such as AAL to covalently bind to the fucose. The lectin resin 1 is incubated with the sample (containing fucose glycoprotein and protein). Fucoose glycoprotein binds to AAL, while non-fucoose glycoprotein or non-glycoprotein remains in the supernatant, thus enriching the fucose glycoprotein. The fucose glycoprotein then binds to an antibody, which in turn binds to the lectin biotin, which in turn binds to HRP for color development.

[0059] Example 2

[0060] Sialic acid glycoprotein enrichment and quantification

[0061] Please see Figure 6 , Figure 1 This diagram illustrates the chemical binding process on the surface of the sialic acid glycoprotein lectin test strip, the binding of fucoglycoprotein to lectin, and the binding process of fucoglycoprotein to antibody in this invention. Figure 6As shown, the test strip surface has groups (aldehyde or ketone groups) that can react with amino groups. Lectins that can enrich sialic acid, such as SNA or MAL-II, are covalently bound to the resin. The test strip is covalently bound to the lectin, which incubates with the sample. This allows sialic acid glycoproteins to bind to the SNA / MAL-II lectin, while non-sialic acid glycoproteins or non-glycoproteins remain in the supernatant, thus enriching the sialic acid glycoproteins. The sialic acid glycoproteins then bind to antibodies, which in turn bind to the lectin biotin, and the biotin binds to HRP for color development.

[0062] Example 3

[0063] Enrichment and quantification of T or Tn antigen glycoproteins

[0064] Please see Figure 7 , Figure 2 This diagram illustrates the chemical binding process on the surface of the T / Tn antigen glycoprotein lectin test strip, the binding of fucose glycoprotein to lectin, and the binding of fucose glycoprotein to antibody in this invention. Figure 7 As shown, the test strip surface has groups (aldehyde or ketone groups) that can react with amino groups, allowing lectins such as PNA or VVL to covalently bind to the resin and enrich T / Tn antigens. The test strip covalently binds to the lectin, which incubates with the sample, causing T / Tn antigen glycoproteins to bind to PNA or VVL, while non-T / Tn antigen glycoproteins remain in the supernatant, thus enriching the T / Tn antigen glycoproteins. The T / Tn antigen glycoproteins then bind to antibodies, which in turn bind to the lectin biotin, which in turn binds to HRP for color development.

[0065] In summary, this invention provides a colorimetric method for fucoidan glycoproteins, a kit utilizing this method, and a method for preparing the kit. First, a liquid sample is dropped onto the surface of the kit. The sample is then introduced into the sample bath and wetted with the test paper. Under lateral flow, the sample flows along the hydrophilic test paper towards the observation window. Glycoproteins containing α1,2-fucose bind to lectin 1, while the remaining sample continues to the next layer of test paper. Glycoproteins containing α1,3-fucose then bind to lectin 2, and the remaining sample flows to the third layer where glycoproteins containing α1,6-fucose bind to lectin 3. All bound fucoidan glycoproteins bind to lectin biotin via their antibodies, and horseradish peroxidase labeling is used for colorimetric detection, achieving quantitative detection of fucoidan glycoproteins in the sample. This method is of great significance for studying the quantitative changes of different linked fucoidan glycoproteins in cells and body fluids during disease.

[0066] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. A method for preparing a fucoidan glycoprotein detection kit, characterized in that, Including the following steps: (1) Select hydrophilic materials containing cellulose as hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3, the lengths of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 increase sequentially, and set one end of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 as lectin binding region respectively; (2) Immerse the lectin binding regions of hydrophilic material 1, hydrophilic material 2 and hydrophilic material 3 in the oxidizing solution and react at room temperature for 1-2 h, then remove them; (3) The lectin-binding region of the hydrophilic material 1 is immersed in the first lectin solution, the lectin-binding region of the hydrophilic material 2 is immersed in the second lectin solution, and the lectin-binding region of the hydrophilic material 3 is immersed in the third lectin solution. All are reacted at room temperature for 6-12 h to obtain the first lectin-binding material, the second lectin-binding material and the third lectin-binding material, respectively. (4) Take out the first lectin binding material, the second lectin binding material and the third lectin binding material, clean them, and vacuum dry the three lectin binding materials at room temperature. (5) Align one end of the first lectin bonding material, the second lectin bonding material and the third lectin bonding material to form an aligned end, and bond the first lectin bonding material, the second lectin bonding material and the third lectin bonding material in order from top to bottom, with the lectin bonding areas all exposed. Then attach a support base plate to the bottom of the third lectin bonding material and attach the fixing material to the support base plate, at which time the fixing material is close to the lectin bonding area. (6) Fix the bottom plate of the box to the bottom of the support base plate, and cover the box cover on the bottom plate to obtain the fucose detection kit. The box cover has a sample injection cell and an observation window. The sample injection cell is located above the alignment end, and the observation window is located above the lectin binding region.

2. The method for preparing a fucoidan glycoprotein detection kit according to claim 1, characterized in that: In step (2), the oxidizing solution is 10-20 mM NaIO3 dissolved in 50-100 mM CH3COONa buffer solution, pH = 4-6.

3. The method for preparing a fucoidan glycoprotein detection kit according to claim 1, characterized in that: In step (3), the first lectin in the first lectin solution is any one or more of UEA-I, PTL-II, and LAA; the second lectin in the second lectin solution is LTL; and the third lectin in the third lectin solution is any one or two of LCH and AAL.

4. The method for preparing a fucose detection kit according to claim 1, characterized in that: In step (4), the cleaning specifically involves immersing the product in 20-30% acetonitrile for 2-4 times, followed by rinsing with deionized water for 3-5 times.

5. A fucoidan glycoprotein detection kit prepared according to any one of claims 1-4, characterized in that: The device includes a cover and a bottom plate, which form a hollow receiving cavity. A sample inlet and an observation window are provided on the cover. A supporting base plate is fixed to the bottom plate. From bottom to top, a third lectin-binding material, a second lectin-binding material, and a first lectin-binding material are sequentially fixed to the supporting base plate. One end of each of these materials is aligned, forming an aligned end, and is located below the sample inlet. The other ends of these materials decrease in size sequentially, forming a stepped shape, so that the lectin-binding area is located below the observation window.

6. The fucoidan glycoprotein detection kit according to claim 5, characterized in that: The thickness of the third lectin-binding material, the second lectin-binding material, and the first lectin-binding material is 0.5-1.0 mm, and the length of the third lectin-binding material, the second lectin-binding material, and the first lectin-binding material decreases by 5-10 mm in sequence.

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