CN115684610A - Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres - Google Patents
Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres Download PDFInfo
- Publication number
- CN115684610A CN115684610A CN202211379037.0A CN202211379037A CN115684610A CN 115684610 A CN115684610 A CN 115684610A CN 202211379037 A CN202211379037 A CN 202211379037A CN 115684610 A CN115684610 A CN 115684610A
- Authority
- CN
- China
- Prior art keywords
- aggregation
- induced emission
- triiodothyronine
- emission polymer
- polymer microspheres
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention discloses a method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres, which dissociates triiodothyronine (T3) combined with thyroid gland-bound globulin by using 8-aniline-1-naphthalenesulfonic acid on an immunochromatographic platform, and avoids interference caused by fluorescent substances in a sample in principle by using special excitation light and emission light bands and a light-emitting mechanism of the aggregation-induced emission polymer microspheres. The excitation light of the selected aggregation-induced emission polymer microsphere is 510nm and the emission light is 615nm. The microsphere can emit orange red light of 615nm under a blue light source of 510nm, and 8-aniline-1-naphthalene sulfonic acid has no fluorescent signal under 510 nm. Therefore, the background fluorescence interference caused by 8-aniline-1-naphthalene sulfonic acid in the dissociating agent is avoided in principle, the interference factors in the detection are reduced, and the reliability of the detection result is improved.
Description
Technical Field
The invention relates to the technical field of in vitro immunodetection, in particular to a method and a kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres.
Background
The thyroid gland is the largest endocrine gland of the human body and can secrete hormones such as thyroxine (T4) and triiodothyronine (T3). These hormones are regulated by Thyroid Stimulating Hormone (TSH) secreted from the pituitary gland, and play an important role in the metabolism of proteins, fats, sugars, vitamins, water and salts, and growth and development of the human body. Thyroid dysfunction is the most common endocrine disorder and is still easy to be missed to diagnose so far. According to the general survey of 4.6 million persons in 11 cities in the united states, about 11% of people have various thyroid disorders that have not been diagnosed. Thyroid disorders can be classified into Hyperthyroidism (Hyperthyroidism) and Hypothyroidism (hypothyroidim).
Triiodothyronine has a molecular weight of 651, approximately 65% is produced directly by the thyroid gland, 35% t4 is formed by deiodination in peripheral tissues. In peripheral blood, 99.6% bind to thyroid-binding globulin (TBG). T3 is the thyroid hormone with the strongest biological activity at present, the biological activity of the T3 is 3-5 times of that of T4, T3 which is not combined with TBG is generally called free T3 clinically, namely free triiodothyronine (FT 3), only the free T3 has metabolic activity, and the combined T3 does not. T3 can be combined with a receptor through a cell membrane to exert physiological effect, so that the T3 is a real active part of the physiological effect of thyroid hormone, and can reflect the functional state of the thyroid and other influences on the function of a human body more accurately.
The immunochromatography reagent is a solid-phase labeling immunoassay technology which organically combines a plurality of methods such as a nano material tracer, an immunoassay technology, a chromatography analysis technology and the like, has the characteristics of simplicity, convenience, time saving, less sample consumption, easy result identification and the like, and can be used for qualitative, semi-quantitative and quantitative detection of a target object in a resource-deficient or non-laboratory environment. Applications include pathogen, drug, hormone and metabolite detection in biomedicine, phytosanitary, veterinary, feed/food and environmental index testing.
Aggregation-induced emission polymer microspheres are prepared by filling Aggregation-induced emission molecules in polystyrene latex microspheres, the quantum yield and the synthesis efficiency of the Aggregation-induced emission polymer microspheres are greatly improved due to limited motion in the filled molecules, the preparation of Aggregation-induced emission polymer microspheres with different excitation light and emission light can be realized according to the difference of the types of the filled Aggregation-induced emission molecules, the Aggregation-induced emission polymer microspheres with different excitation light and emission light are selected according to the actual detection requirements, and the problem of interference of fluorescent substances in a sample to be detected can be well solved due to the unique Aggregation-induced emission (AIE) mechanism of the Aggregation-induced emission molecules.
Currently, 8-aniline-1-naphthalenesulfonic Acid (ANS) is generally used to dissociate triiodothyronine (T3) molecules bound to thyroid-associated globulin, and a chemiluminescence scheme is used to quantify the amount of triiodothyronine in serum through the interaction of specific hapten and antibody. In the traditional fluorescence immunochromatography, most of used tracers are time-resolved fluorescent microspheres and quantum dots, the excitation light of the tracers is 360-400nm, the excitation light of 8-aniline-1-naphthalenesulfonic acid required for dissociating thyroid-associated globulin-bound triiodothyronine (T3) molecules is also in the ultraviolet band, and the emitted light of the tracers is 470-480nm and presents blue light. Therefore, the fluorescent microspheres and the quantum dots have larger background interference on the fluorescent signals of the time-resolved fluorescent microspheres and the quantum dots.
Disclosure of Invention
The invention aims to solve the technical problem of providing a method for detecting triiodothyronine which is not easily influenced by the exciting light of 8-aniline-1-naphthalene sulfonic acid
In order to solve the technical problems, the invention aims to realize the following technical scheme: the method for detecting triiodothyronine based on the aggregation-induced emission polymer microspheres comprises the following steps:
s1, preparing a T3 dissociating agent;
s2, adding the serum to be detected and the T3 dissociation agent into a centrifugal tube, oscillating and uniformly mixing to form a sample diluent to be detected, then dropwise adding the sample diluent to be detected into a sample adding hole of the T3 immunochromatography reagent strip, and judging by using a dry fluorescence immunoassay analyzer after 15 min;
wherein, the T3 immunochromatography reagent strip is provided with a compound of an aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and a chicken egg yolk antibody.
Preferably, the preparation of the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody comprises the following steps:
a1, activation: adding 100 μ L of 1% aggregation-induced emission polymer microspheres into 400 μ L of 10-50mM MES buffer solution with pH of 5.0-7.0, adding 100-500 μ g EDC and 150-750 μ g NHS activator, and activating in 37 deg.C water bath;
a2, centrifugal redissolution: centrifuging at 18000-30000g for 10min after activation, removing supernatant, and re-dissolving fluorescent microsphere with 200-400 μ L10-50mM pH7.0-8.0 boric acid buffer solution;
a3, adding an antibody: 5-50 mug of T3 specific monoclonal antibody and 10-30 mug of chicken egg yolk antibody are respectively added, and the reaction is carried out in water bath at 37 ℃ for 120 minutes.
Preferably, the preparation of the T3 immunochromatographic reagent strip comprises the following steps:
b1, preparing a detection line and a quality control line: the detection line is that triiodothyronine is coupled with bovine serum albumin antigen and is diluted by boric acid buffer solution and then is scribed on a nitrocellulose membrane; the quality control line is drawn on a nitrocellulose membrane after the goat anti-chicken antibody is diluted by using a boric acid buffer solution;
b2, preparing a bonding pad: diluting the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody by using a diluent, and spraying the diluted compound on a glass fiber membrane;
b3, preparing a sample pad: placing the whatman-MF1 blood filtering membrane in a sample pad treatment solution for soaking, and then spreading and drying;
and B4, assembling.
Preferably, in the step B1, the concentration of the boric acid buffer solution is 10-50mM, and the concentration of the triiodothyronine coupled with bovine serum albumin antigen after dilution is 0.1-1.0mg/mL.
Preferably, the concentration of the diluted goat anti-chicken antibody is 0.01-0.5mg/mL.
Preferably, in the step B2, 5 to 10. Mu.L of the aggregation inducing luminescent polymer microspheres are collectively labeled with the complexes of the T3-specific monoclonal antibody and the chicken egg yolk antibody, the volume of the dilution is 290. Mu.L, and the dilution is a mixed solution of trehalose at a final concentration of 5 to 20%, 10 to 50mM Tris, 2 to 10% BSA and 0.5 to 2% Tw-20.
Preferably, the sample pad treatment solution includes 10-100mM boric acid-borax buffer solution, 2-5% trehalose, 0.5-2.0% casein and 0.5-1.5% Tw-20%.
Preferably, the T3 cleavage agent comprises the following components: purified water, tris, naCl, proclin300 and a dissociating agent ANS.
Preferably, the T3 cleavage agent comprises the following components: 1000mL of purified water, 0.605g of Tris, 9g of NaC1, 0.5mL of proclinin 300 and 1-3g of a dissociating agent ANS.
8-aniline-1-naphthalenesulfonic acid was used on the immunochromatographic platform to dissociate triiodothyronine (T3) bound to thyroid-bound globulin. And interference caused by fluorescent substances in a sample is avoided in principle by using special excitation light and emission light bands and a light-emitting mechanism of the aggregation-induced emission polymer microsphere. The excitation light of the selected aggregation-induced emission polymer microsphere is 510nm and the emission light is 615nm. The microsphere can emit orange red light of 615nm under a blue light source of 510nm, and 8-aniline-1-naphthalene sulfonic acid has no fluorescent signal under 510 nm. Therefore, the background fluorescence interference caused by 8-aniline-1-naphthalene sulfonic acid in the dissociating agent is avoided in principle, the interference factors in the detection are reduced, and the reliability of the detection result is improved.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the description of the embodiments are briefly introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings based on these drawings without creative efforts.
Fig. 1 is a structural diagram of an immunochromatographic test strip provided in an embodiment of the present invention;
FIG. 2 is a graph of excitation and emission spectra of aggregation-induced emission polymeric microspheres according to an embodiment of the present invention;
FIG. 3 is the fluorescence signal intensity of the T and C lines and the T3 content data of the sample according to the application example of the present invention;
FIG. 4 is a regression line graph of clinical relevance of an example of an application of the present invention.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the embodiments of the present invention, and it should be apparent that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be obtained by a person skilled in the art without making any creative effort based on the embodiments in the present invention, belong to the protection scope of the present invention.
It will be understood that the terms "comprises" and/or "comprising," when used in this specification and the appended claims, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof.
It is also to be understood that the terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used in this specification and the appended claims, the singular forms "a", "an", and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise.
It should be further understood that the term "and/or" as used in this specification and the appended claims refers to any and all possible combinations of one or more of the associated listed items and includes such combinations.
Furthermore, the terms "substantially", "essentially", and the like are intended to indicate that the relative terms are not required to be absolutely exact, but may have some deviation. For example: "substantially equal" does not mean absolute equality, but it is difficult to achieve absolute equality in actual production and operation, and certain deviations generally exist. Thus, in addition to absolute equality, "substantially equal" also includes the above-described case where there is some deviation. In this case, unless otherwise specified, terms such as "substantially", and the like are used in a similar manner to those described above.
In the description of the present invention, it should also be noted that, unless otherwise explicitly specified or limited, the terms "disposed," "mounted," "connected," and "connected" are to be construed broadly and may, for example, be fixedly connected, detachably connected, or integrally connected; they may be connected directly or indirectly through intervening media, or they may be interconnected between two elements. The specific meanings of the above terms in the present invention can be understood in specific cases to those skilled in the art.
In order to reduce background interference of excitation light of 8-aniline-1-naphthalenesulfonic acid on detecting triiodothyronine when 8-aniline-1-naphthalenesulfonic acid is used for dissociating triiodothyronine (T3) molecules bound with thyroid gland-bound globulin and realizing quantification of the triiodothyronine in serum through interaction of specific hapten and antibody by adopting a chemiluminescence scheme, the invention provides a method for detecting the triiodothyronine based on aggregation-induced luminescent polymer microspheres.
The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres provided by the invention comprises the following steps:
s1, preparing a T3 dissociating agent;
s2, adding the serum to be detected and the T3 dissociation agent into a centrifuge tube, oscillating and uniformly mixing to form a sample diluent to be detected, then dropwise adding the sample diluent to be detected into a sample adding hole of the T3 immunochromatography reagent strip, and after 15min, using a dry fluorescence immunoassay analyzer to interpret;
wherein, the T3 immunochromatography reagent strip is provided with a compound of an aggregation induced emission polymer microsphere labeled T3 specific monoclonal antibody and a chicken egg yolk antibody.
The T3 dissociation agent comprises the following components: purified water, tris, naCl, proclin300 and a dissociating agent ANS. Specifically, 1000mL of purified water, 0.605g of Tris, 9g of NaC1, 0.5mL of Proclin300, and 1-3g of a dissociating agent ANS.
The preparation of the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody comprises the following steps:
a1, activation: adding 100 mu L of 1% aggregation-induced emission polymer microspheres into 400 mu L of 10-50mM MES buffer solution with pH value of 5.0-7.0 in percentage by mass, adding 100-500 mu g of EDC and 150-750 mu g of NHS activating agent, and performing water bath activation at 37 ℃;
a2, centrifugal redissolution: centrifuging at 18000-30000g for 10min after activation, removing supernatant, and re-dissolving fluorescent microsphere with 200-400 μ L10-50mM pH7.0-8.0 boric acid buffer solution;
a3, adding an antibody: 5-50 mug of T3 specific monoclonal antibody and 10-30 mug of chicken egg yolk antibody are respectively added, and the reaction is carried out in water bath at 37 ℃ for 120 minutes.
On the other hand, the preparation of the T3 immunochromatography reagent strip comprises the following steps:
b1, preparing a detection line and a quality control line: the detection line is that triiodothyronine is coupled with bovine serum albumin antigen and is diluted by boric acid buffer solution and then is scribed on a nitrocellulose membrane; the quality control line is formed by diluting goat anti-chicken antibody with boric acid buffer solution and then scribing on a nitrocellulose membrane;
b2, preparing a bonding pad: diluting the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody by using a diluent, and spraying the diluted compound on a glass fiber membrane;
b3, preparing a sample pad: placing the whatman-MF1 blood filtering membrane in a sample pad treatment solution for soaking, and then spreading and drying;
and B4, assembling.
In the present invention, there are two kinds of microsphere complexes on the conjugate pad, one is a complex bound with a T3 specific monoclonal antibody, which can bind to T3-BSA on the T line, but if there is T3 in the sample, the complex will first contact T3 in the sample and will first bind to T3 in the sample to form a T3-T3 specific antibody-microsphere complex, which cannot bind to T3-BSA on the T line because the sites of the antibody are occupied. So the number of microspheres on the T line is more or less (the number of microspheres is converted into the intensity of a fluorescence signal by an instrument). The signal is strongest in the absence of T3 in the sample and decreases in the presence of T3.
Another microsphere compound is prepared by combining chicken yolk antibody-microsphere with sheep anti-chicken specific binding on C line. The main function of the C line is to ensure the stability of the T/C value.
The sample pad can filter the sample and simultaneously enable the sample to flow upwards; and the sample pad treatment solution contains a surfactant and a protein substance, so that the immunoreaction on the reagent strip can be ensured to be stably carried out in the chromatography process.
In addition, in the step B1, the concentration of the boric acid buffer solution is 10-50mM, the concentration of triiodothyronine coupled with bovine serum albumin antigen after dilution is 0.1-1.0mg/mL, and the concentration of goat anti-chicken antibody after dilution is 0.01-0.5mg/mL.
In the step B2, 5 to 10. Mu.L of the aggregation inducing luminescent polymer microsphere labeled complex of the T3-specific monoclonal antibody and the chicken egg yolk antibody was taken in total, the volume of the diluent was 290. Mu.L, and the diluent was a mixed solution of trehalose at a final concentration of 5 to 20%, 10 to 50mM (0.121 to 0.6%) Tris, 2 to 10% BSA and 0.5 to 2% Tw-20.
In step B3, the sample pad treatment solution comprises 10-100mM (0.06-0.1%) boric acid-borax buffer solution, 2-5% trehalose, 0.5-2.0% casein and 0.5-1.5% wt% Tw-20.
8-aniline-1-naphthalenesulfonic acid was used on the immunochromatographic platform to dissociate triiodothyronine (T3) bound to thyroid-bound globulin. And interference caused by fluorescent substances in a sample is avoided in principle by using special excitation light and emission light wave bands and a light-emitting mechanism of the aggregation-induced emission polymer microsphere. The excitation light of the selected aggregation-induced emission polymer microsphere is 510nm and the emission light is 615nm. The microsphere can emit orange red light of 615nm under a blue light source of 510nm, and 8-aniline-1-naphthalene sulfonic acid has no fluorescent signal under 510 nm. Therefore, the background fluorescence interference caused by 8-aniline-1-naphthalene sulfonic acid in the dissociating agent is avoided in principle, the interference factors in the detection are reduced, and the reliability of the detection result is improved.
The following specific description is given with reference to the following examples and application examples:
examples
S1, preparing a T3 dissociating agent;
s2, adding the serum to be detected and the T3 dissociation agent into a centrifugal tube, oscillating and uniformly mixing to form a sample diluent to be detected, then dropwise adding the sample diluent to be detected into a sample adding hole of the T3 immunochromatography reagent strip, and judging by using a dry fluorescence immunoassay analyzer after 15 min;
wherein, the T3 immunochromatography reagent strip is provided with a compound of an aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and a chicken egg yolk antibody.
The T3 dissociation agent specifically comprises the following components: 1000mL of purified water, 0.605g of Tris, 9g of NaC1, 0.5mL of Proclin300 and 2g of dissociating agent ANS.
The preparation method of the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody comprises the following steps:
a1, activation: 100 μ L of 1% aggregation-induced emission polymer microspheres are activated in a water bath at 37 ℃ after being added with 400 μ L of 30mM MES and pH5.0-7.0 buffer solution and 100-500 μ g of EDC and 450 μ g of NHS activator according to mass percentage;
a2, centrifugal redissolution: centrifuging at 18000-30000g for 10min after activation, removing supernatant, and re-dissolving fluorescent microsphere with 200-400 μ L10-50mM pH7.0-8.0 boric acid buffer solution;
a3, adding an antibody: 5-50 mug of T3 specific monoclonal antibody and 10-30 mug of chicken egg yolk antibody are respectively added, and the reaction is carried out in water bath at 37 ℃ for 120 minutes.
On the other hand, the preparation of the T3 immunochromatography reagent strip comprises the following steps:
b1, preparing a detection line and a quality control line: the detection line is that triiodothyronine is coupled with bovine serum albumin antigen and is diluted by boric acid buffer solution and then is scribed on a nitrocellulose membrane; the quality control line is drawn on a nitrocellulose membrane after the goat anti-chicken antibody is diluted by using a boric acid buffer solution;
b2, preparing a bonding pad: diluting the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody by using a diluent, and spraying the diluted compound on a glass fiber membrane;
b3, preparing a sample pad: placing the whatman-MF1 blood filtering membrane in a sample pad treatment solution for soaking, and then spreading and drying;
and B4, assembling the semi-finished products prepared in the steps B1-B3 on a PVC (polyvinyl chloride) bottom plate according to the mode of the figure 1, and using a strip cutting machine to cut reagent strips with the length of 3.9mm into corresponding reagent card shells, thus finishing the preparation of the corresponding reagent cards.
In the step B1, the concentration of the boric acid buffer solution is 10-50mM, the concentration of triiodothyronine coupled with bovine serum albumin antigen after dilution is 0.1-1.0mg/mL, and the concentration of goat anti-chicken antibody after dilution is 0.01-0.5mg/mL.
In step B2, a total of 5-10. Mu.L of the aggregation inducing luminescent polymer microspheres labeled with a complex of the T3-specific monoclonal antibody and the chicken egg yolk antibody was taken, and the diluted solution was 290. Mu.L of trehalose at a final concentration of 5-20%, 10-50mM Tris, 2-10% BSA and 0.5-2% Tw-20 as a mixed solution.
In step B3, the sample pad treatment solution comprises 10-100mM boric acid-borax buffer solution, 2-5% trehalose, 0.5-2.0% casein and 0.5-1.5% wt% Tw-20.
Application example
The reagent strip prepared in the embodiment is used for testing 40 clinical serum samples, the concentration value of the T3 content in the 40 clinical serum samples is determined by Roche T3 kit detection, and the performance of the reagent strip and the correlation of the clinical samples are evaluated by comparing the T/C value of the reagent strip with the concentration value tested by the Roche T3 kit.
And (3) detection flow: 100 mu L of serum to be detected is taken by a pipette, added into 200 mu L of dissociating agent and mixed evenly for 30 seconds, so that ANS can fully decompose T3 combined with TBG. Then 80. Mu.L of the mixed liquid is slowly added into the sample adding hole of the detection card, and timing is started. When the reaction time is 15 minutes, the detection card is immediately inserted into a fluorescence immunoassay quantitative analyzer, the fluorescence signal intensity of the T line and the C line is read, and detailed data are shown in figure 3; the performance of the agents and their clinical relevance are shown in FIG. 4.
As can be seen from FIG. 4, the reagent card has good linearity in the range of T3 concentration of 0.3-10nmol/L, and the clinical relevance R 2 =0.9608, the performance was essentially identical to that of the T3 kit from roche.
While the invention has been described with reference to specific embodiments, the invention is not limited thereto, and various equivalent modifications and substitutions can be easily made by those skilled in the art within the technical scope of the invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the claims.
Claims (10)
1. A method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres is characterized by comprising the following steps:
s1, preparing a T3 dissociating agent;
s2, adding the serum to be detected and the T3 dissociation agent into a centrifugal tube, oscillating and uniformly mixing to form a sample diluent to be detected, then dropwise adding the sample diluent to be detected into a sample adding hole of the T3 immunochromatography reagent strip, and judging by using a dry fluorescence immunoassay analyzer after 15 min;
wherein, the T3 immunochromatography reagent strip is provided with a compound of an aggregation induced emission polymer microsphere labeled T3 specific monoclonal antibody and a chicken egg yolk antibody.
2. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres as claimed in claim 1, wherein the preparation of the complex of the aggregation-induced emission polymer microspheres labeled with T3 specific monoclonal antibody and chicken egg yolk antibody comprises the following steps:
a1, activation: adding 100 μ L of 1% aggregation-induced emission polymer microspheres into 400 μ L of 10-50mM MES buffer solution with pH of 5.0-7.0, adding 100-500 μ g EDC and 150-750 μ g NHS activator, and activating in 37 deg.C water bath;
a2, centrifugal redissolution: centrifuging at 18000-30000g for 10min after activation, removing supernatant, and re-dissolving fluorescent microsphere with 200-400 μ L10-50mM pH7.0-8.0 boric acid buffer solution;
a3, adding an antibody: 5-50 mug of T3 specific monoclonal antibody and 10-30 mug of chicken egg yolk antibody are respectively added, and the reaction is carried out in water bath at 37 ℃ for 120 minutes.
3. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres according to claim 2, wherein the preparation of the T3 immunochromatographic reagent strip comprises the following steps:
b1, preparing a detection line and a quality control line: the detection line is that triiodothyronine is coupled with bovine serum albumin antigen and is diluted by boric acid buffer solution and then is scribed on a nitrocellulose membrane; the quality control line is formed by diluting goat anti-chicken antibody with boric acid buffer solution and then scribing on a nitrocellulose membrane;
b2, preparing a bonding pad: diluting the compound of the aggregation-induced emission polymer microsphere labeled T3 specific monoclonal antibody and the chicken egg yolk antibody by using a diluent, and spraying the diluted compound on a glass fiber membrane;
b3, preparing a sample pad: placing the whatman-MF1 blood filtering membrane in a sample pad treatment solution, soaking, then spreading and drying;
and B4, assembling.
4. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres as claimed in claim 3, wherein: in the step B1, the concentration of the boric acid buffer solution is 10-50mM, and the concentration of the triiodothyronine coupled with the bovine serum albumin antigen after dilution is 0.1-1.0mg/mL.
5. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres as claimed in claim 3, wherein: the concentration of the goat anti-chicken antibody after dilution is 0.01-0.5mg/mL.
6. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres as claimed in claim 3, wherein: in the step B2, 5-10 mu L of aggregation induced emission polymer microsphere is taken together to mark the compound of the T3 specific monoclonal antibody and the chicken egg yolk antibody, and the volume of the diluent is 290 mu L ， And the diluent is a mixed solution of trehalose at a final concentration of 5-20%, 10-50mM Tris, 2-10% BSA and 0.5-2% Tw-20%.
7. The method of claim 3, wherein the sample pad treatment solution comprises 10-100mM boric acid-borax buffer solution, 2-5% trehalose, 0.5-2.0% casein and 0.5-1.5%.
8. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres as claimed in claim 1, wherein the T3 dissociation agent comprises the following components: purified water, tris, naCl, proclin300 and a dissociating agent ANS.
9. The method for detecting triiodothyronine based on aggregation-induced emission polymer microspheres of claim 8, wherein the T3 dissociation agent comprises the following components: 1000mL of purified water, 0.605g of Tris, 9g of NaC1, 0.5mL of proclinin 300 and 1-3g of a dissociating agent ANS.
10. A kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres is characterized in that: comprising the T3 dissociation agent of any one of claims 1-9 and a T3 immunochromatographic reagent strip.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211379037.0A CN115684610A (en) | 2022-11-04 | 2022-11-04 | Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211379037.0A CN115684610A (en) | 2022-11-04 | 2022-11-04 | Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN115684610A true CN115684610A (en) | 2023-02-03 |
Family
ID=85049874
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211379037.0A Pending CN115684610A (en) | 2022-11-04 | 2022-11-04 | Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN115684610A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117169519A (en) * | 2023-10-26 | 2023-12-05 | 艾康生物技术(杭州)有限公司 | Dissociation agent and kit for detecting TT3 and/or TT4 in sample |
-
2022
- 2022-11-04 CN CN202211379037.0A patent/CN115684610A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117169519A (en) * | 2023-10-26 | 2023-12-05 | 艾康生物技术(杭州)有限公司 | Dissociation agent and kit for detecting TT3 and/or TT4 in sample |
| CN117169519B (en) * | 2023-10-26 | 2024-01-30 | 艾康生物技术(杭州)有限公司 | Dissociation agent and kit for detecting TT3 and/or TT4 in sample |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| RU2608656C2 (en) | Magnetic particles associated with streptavidin and method of production thereof | |
| WO2020199501A1 (en) | Quick detection method for food allergens based on quantum dot fluorescence | |
| CN106153927A (en) | A kind of fast quantification detects time-resolved fluoroimmunoassay chromatography reagent and the preparation method of cTnI, CKMB, Myo simultaneously | |
| CN1928561A (en) | Photon crystal micro-ball based multicomponent immunity detection method | |
| EP3904878A1 (en) | Immunochromatography strip for pregnancy diagnosis with multiple test lines, and pregnancy diagnosis kit comprising same | |
| CN107918022B (en) | cTnI detection kit and use method thereof | |
| CN115684610A (en) | Method and kit for detecting triiodothyronine based on aggregation-induced emission polymer microspheres | |
| CN110873800A (en) | Glycosylated hemoglobin immunochromatographic test strip and preparation method and kit thereof | |
| CN110927394A (en) | BNP, NT-proBNP and sST2 combined detection reagent strip and preparation method thereof | |
| CN113125696B (en) | Estradiol homogeneous chemiluminescence detection kit and application thereof | |
| CN210269868U (en) | Joint detection mechanism and female fertility detection device | |
| CN104387467A (en) | Detection kit and detection paper for Beta-adrenergic receptor stimulant multiresidue detection | |
| KR102346178B1 (en) | Strips, kits for immunochromatography and competitive immunochromatographic assays using the same | |
| JPS63127160A (en) | Detection of specific protein | |
| CN104311659A (en) | Multi-cluster antigen and wide-spectrum specific antibody of beta-adrenoceptor agonists and application of multi-cluster antigen and wide-spectrum specific antibody | |
| CN106124777A (en) | Acridine labelling conjugate and preparation method thereof, chemical luminescence reagent kit | |
| JP2013527467A5 (en) | ||
| CN113125697B (en) | Testosterone homogeneous phase chemiluminescence detection kit and application thereof | |
| CN106124771A (en) | A kind of step homogeneous cTnT detection kit and application thereof | |
| CN113109325A (en) | Pepsinogen I enzymatic chemiluminescence detection kit and preparation method and application thereof | |
| JPS58122459A (en) | Measuring method utilizing association of enzyme | |
| CN110579596A (en) | Multi-joint detection immunochromatography reagent card, application thereof and multi-joint detection immunochromatography kit | |
| CN111381026A (en) | Multiple detection immunoassay reagent, preparation method, kit, system and application thereof | |
| EP2230516B1 (en) | Method of immunoassaying specimen using aggregation reaction of microparticles and assay kit | |
| CN219434848U (en) | Quantitative joint inspection immunochromatography kit |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |