CN109142743B - Multiple detection kit for self-immune liver antibody - Google Patents
Multiple detection kit for self-immune liver antibody Download PDFInfo
- Publication number
- CN109142743B CN109142743B CN201810823210.9A CN201810823210A CN109142743B CN 109142743 B CN109142743 B CN 109142743B CN 201810823210 A CN201810823210 A CN 201810823210A CN 109142743 B CN109142743 B CN 109142743B
- Authority
- CN
- China
- Prior art keywords
- microbeads
- microbead
- autoimmune liver
- beads
- detection
- 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.)
- Active
Links
Classifications
-
- 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
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6854—Immunoglobulins
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Food Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Biotechnology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a multiple detection kit for an autoimmune liver antibody, which comprises: a plurality of sealed reagent bottles which are respectively filled with the composite microbead suspension, the fluorescent marker, the sample diluent and the washing solution, and a kit which separates and collectively packages the reagent bottles; the micro-beads are polystyrene micro-beads dyed by different fluorescent dyes and coded; the composite microbead suspension comprises detection microbeads, standard microbeads, reference microbeads and blank microbeads, wherein the detection microbeads are different autoimmune liver disease antigens respectively coated on the surfaces of the microbeads with different codes; the standard microbeads comprise at least three microbeads combined with human IgG with different concentration gradients; the reference beads are anti-human IgG-bound beads; blank beads are beads to which no antigen has been added. The invention can simultaneously detect a plurality of self-immune liver antibodies in a human serum sample, provides an effective laboratory detection means for patients with autoimmune liver diseases, and has important significance for diagnosis and further treatment.
Description
Technical Field
The invention relates to the field of detection of autoimmune liver disease antibodies, in particular to a kit capable of detecting multiple autoimmune liver antibodies at one time.
Background
Generally, humans do not produce autoantibodies. The primary function of the immune system is to distinguish foreign antigens (e.g., infectious antigens) from self-tissues. Immune confusion occurs when a person's immune system produces antibodies (i.e., autoantibodies) that are potentially destructive to self-antigens. Most autoimmune disorders can be classified as non-organ specific (systemic) or organ specific.
Autoimmune liver disease, a disease with unknown pathogenesis, is associated with many diseases, and clinical diagnosis is difficult because the initial clinical symptoms are similar to those of viral hepatitis. Therefore, accurately and timely monitoring the specificity of autoantibodies in autoimmune liver diseases is undoubtedly of great significance for diagnosis and further treatment.
The autoimmune liver disease antibody comprises immunoglobulin IgG antibodies of various antigens such as Ro-52(SSA 52kDa), mitochondria (AMA), anti-CENP B, AMA-M2, Actin, LKM-1, Sp100, gp210, Nup62, PML, LC-1 and SAL/LP).
The Ro-52 antibody (Ro-52) anti-Ro 52 antibody has no disease specificity, and can be detected in myositis, systemic sclerosis, other collagenosis, neonatal lupus erythematosus, primary biliary cirrhosis, autoimmune hepatitis and viral hepatitis.
Anti-mitochondrial antibody-M2: AMA-M2 is the most sensitive and specific diagnostic marker for Primary Biliary Cirrhosis (PBC). High titers of AMA-M2 antibodies are important for the diagnosis of PBC and for predicting whether PBC patients will develop symptoms of liver dysfunction or cholestasis. Antibodies to AMA-M2 may also be detected in patients with autoimmune hepatitis, progressive systemic sclerosis, sjogren's syndrome, and who overlap with PBC. Low-concentration AMA-M2 antibody can be detected by chronic HCV and systemic lupus erythematosus.
Promyelocytic protein antibody (PML) PML antibodies were predominantly present in patients positive for Sp100 antibody. Both have the same sensitivity and specificity. PBC patients who appeared positive for PML antibody and Sp100 antibody had a faster progression with poorer prognosis. The high specificity of PML and Sp100 improves the detection rate of PBC patients, and is an important index for early diagnosis of PBC.
The nuclear granule protein antibody (Sp100) has a small proportion of PBC and Sp100 antibody is detectable in patients with autoimmune hepatitis (AIH).
Anti-gp 210 antibody (gp210) anti-gp 210 antibody has high sensitivity and high specificity to PBC, and detection in combination with Sp100 and AMA-M2 can improve the detection rate of PBC patients and increase specificity to PBC diagnosis. gp210, Sp100 is of great significance for PBC diagnosis, especially for patients who are AMA-M2 negative.
The liver and kidney microsome antibody (LKM-1) anti-liver and kidney microsome antibody-1 is a marker antibody of II type AIH, and plays an important role in diagnosis and differential diagnosis. The development of molecular biology technology provides a very useful experimental means for identifying the self-target antigen of autoimmune diseases. In addition to autoimmune hepatitis type II, LKM-1 is also present in the serum of a small number of hepatitis C patients. LKM-1 can be detected in 1% of adult AIH patients, but LKM-1 is more positive in pediatric AIH patients. LKM-1 can also be detected in 1-2% of HCV patients.
The liver solute antibody (LC-1) LC-1 has certain sensitivity and 100% specificity on the diagnosis of AIH, and can improve the detection rate of AIH patients when being detected together with SLA/LP. The positive rate of the soluble liver-hepato-pancreatic antibody (SLA/LP) SLA/LP in the patients with AIH is 30%, but the specificity is 100%, and if there are corresponding clinical symptoms, only SLA/LP can basically diagnose AIH.
Smooth Muscle Antibody (SMA): autoimmune hepatitis can develop high titers of SMA, often with ANA. Anti-actin antibodies, which are commonly found in AIH patients, are also found in various liver diseases or rheumatic diseases, etc. The simultaneous occurrence (positive) of SMA and ANA with high titer is the most important reference index for diagnosing type I AIH, the positive rate is as high as 92.2%, and the antibody has high sensitivity but poor specificity. The single autoantibody test cannot diagnose AIH and needs to be combined with other clinical indicators to diagnose.
Soluble liver antigen/hepatic pancreatic antigen (SLA/LP) antibody SLA/LP antibody is the most specific diagnostic marker for AIH. Although the positive rate is only 10-30%, the positive prediction value is almost 100%. Antibody positivity essentially diagnoses AIH if the corresponding clinical symptoms appear. anti-SLA/LP is highly specific for AIH and can be detected in sera that are negative for low titers of ANA or other autoantibodies. Therefore, the antibody detection is carried out on the ANA, SMA and LKM-l antibody negative or low-titer liver disease patients, the cryptogenic chronic liver disease patients can be reclassified, the accuracy of AIH diagnosis is improved, missed diagnosis and misdiagnosis are reduced, and the atypical AIH patients are effectively treated in time. But the detection rate of anti-SLA/LP is low. Kanzler et al reported only 11% to 32.5% of AIH patients to be positive for anti-SLA/LP. In view of this situation, there is a need to find more sensitive detection methods to increase the detection rate of anti-SLA/LP for clinical diagnosis.
anti-Centromere (Centromere) antibodies are mainly found in CREST syndrome, i.e. calcification, raynaud's phenomenon, esophageal dyskinesia, scleroderma and telangiectasia, with a detection rate of 38-80%. Diffuse scleroderma (detection rate of about 20%) and Primary Biliary Cirrhosis (PBC) are also seen. ACA-positive is often accompanied by other PBC-associated autoantibodies in PBC patients, with about 20% AMA-positive being accompanied by ACA-positive in PBC patients. ACA-associated anti-nuclear point antibodies (SP100, PML, etc.) and anti-nuclear envelope protein antibodies (gp210, p62) are also found. The simultaneous presence of multiple PBC-associated autoantibodies may increase specificity of PBC diagnosis.
An anti-nucleoporin (nuclear pore protein) p62 antibody, the target antigen of which is a 62kD transmembrane protein located on the nuclear pore complex. The anti-p 62 antibody is another high specificity autoantibody to PBC, which is not detected in other liver diseases or autoimmune diseases, and the sensitivity is 23% -32%. Anti-gp 210 antibodies and anti-p 62 antibodies tend to be independent of each other and generally not positive at the same time.
In recent years, there have been various methods for detecting antibodies against autoimmune liver diseases, such as indirect fluorescence, Uhur's gel diffusion, blood coagulation, radioimmunoassay or enzyme assay. However, the above method cannot simultaneously detect a plurality of autoimmune liver disease antibodies in an immunoassay.
Disclosure of Invention
The invention aims to provide a multiple detection kit for the self-immune liver antibody, so as to solve the defects in the background technology.
The invention is realized by the following technical scheme:
a multiple-item test kit for an autoimmune liver antibody, the kit comprising:
(1) a plurality of sealed reagent bottles respectively filled with the composite microbead suspension, the fluorescent marker, the sample diluent and the washing solution, and (2) a kit for separating and packaging the reagent bottles in a centralized manner; the microbeads are polystyrene microspheres with uniform size, and the polystyrene microspheres are dyed by different fluorescent dyes, so that microbeads with different codes (such as No. 1-100) are obtained; the composite microbead suspension comprises detection microbeads, standard microbeads, reference microbeads and blank microbeads, and specifically comprises the following components:
the detection beads are formed by respectively coating different autoimmune liver disease antigens on the surfaces of beads with different codes;
the standard microbeads comprise at least three microbeads with different concentration gradients of human IgG bound thereto;
the reference beads are anti-human IgG-bound beads;
the blank beads are beads to which no antigen is added.
Preferably, the beads have a diameter of 5 to 7 microns, most preferably 5.6 microns.
The autoimmune liver disease antigens include: SSA 52, anti-CENP B, AMA-M2, Actin, LKM-1, Sp100, gp210, Nup62, PML, LC-1, and SAL/LP.
Preferably, the fluorescent marker is phycoerythrin-labeled goat anti-human IgG (specific gamma chain).
The sample diluent is a phosphate-containing buffer. Preferably, the composition and content of each component in the sample diluent are as follows:
the washing solution has the same composition as the sample diluent and has a concentration 8 to 10 times that of the sample diluent.
As a preferable technical scheme, the multiple detection kit for the autoimmune liver antibody of the present invention further comprises a reagent bottle containing a quality control product, wherein the quality control product comprises a positive quality control product and a negative quality control product, the positive quality control product is human mixed serum containing an antibody corresponding to the autoimmune liver disease antigen contained in the detection beads, and the negative quality control product is human negative serum negative to the antibody corresponding to the autoimmune liver disease antigen. The concentration range of the positive quality control has batch-to-batch specificity, i.e., each batch of reagent has its own concentration range.
The preparation method of the composite microbead suspension comprises the following steps:
and respectively coating different autoimmune liver disease antigens by using microbeads with different codes, mixing to prepare detection microbeads, transferring the detection microbeads, the standard microbeads, the reference microbeads and the blank microbeads to a sample diluent, and mixing to obtain the composite microbead suspension.
The detection microbead is prepared by the following method: preparing the micro-bead activating solution, adding any one autoimmune liver disease antigen according to the proportion that each milliliter of micro-bead contains 10-100ug of antigen, and fully mixing. Coating different autoimmune liver disease antigens requires the use of different encoded microbeads.
The microbead activating solution is prepared by adding microbeads of 1000 unit volumes into a reaction tube, and adding EDC (1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride) of 5-50 unit volumes and Sulfo-NHS (N-hydroxy thiosuccinimide) of 5-50 unit volumes for activation. Different test items coat different antigens and require the use of different encoded microbeads.
The standard microbead is prepared by the following method: the bead activating solution was prepared, and then human IgG (immunoglobulin G) was added in a proportion of 2-18ug per ml of beads, followed by thorough mixing. The content of the human IgG in the three or more standard microbeads is uniformly and sequentially increased within the range of 2-18 ug. For example, when 3 kinds of standard microbeads are used, the concentration of human IgG is 2-5ug/ml, 6-10ug/ml, 12-18ug/ml, respectively.
The reference microbead is prepared by the following method: preparing a microbead activating solution, adding anti-human IgG into each milliliter of microbead according to the proportion of 8-15ug of anti-human IgG, and fully mixing.
The blank microbead is the microbead activating solution without any antigen added.
The multiple detection kit for the autoimmune liver antibody has the innovation point that the detection of multiple autoimmune liver disease antibodies can be carried out on one human serum sample in the same hole at the same time. Autoimmune liver disease, a disease with unknown pathogenesis, is associated with many diseases, and clinical diagnosis is difficult because the initial clinical symptoms are similar to those of viral hepatitis. Therefore, accurately and timely monitoring the specificity of autoantibodies in autoimmune liver diseases is undoubtedly of great significance for diagnosis and further treatment. The invention undoubtedly provides an effective laboratory detection means for patients with autoimmune liver diseases.
Detailed Description
The invention is illustrated below by means of specific examples, without being restricted thereto.
Example 1:
the preparation process of the multiple detection kit for the autoimmune liver antibody comprises the following steps:
1. preparation of sample dilutions
The formulation for 8L was as follows:
| numbering | Name of reagent | Concentration of |
| 1 | NaCl | 8.0g/L |
| 2 | Na2HPO4·12H2O | 2.9g/L |
| 3 | KCl | 2.4g/L |
| 4 | KH2PO4 | 2.4g/L |
| 5 | Tris | 0.3g/L |
| 6 | NaN3 | 0.5g/L |
2. Activation of Microbeads
1ml of 5.6-micron microbeads (polystyrene microspheres from Luminex, USA) are added into a 1.5ml centrifuge tube, and 5-50ul of EDC (1-ethyl- (3-dimethylaminopropyl) carbonyl diimine hydrochloride) and 5-50 unit volume of Sulfo-NHS (N-hydroxy thiosuccinimide) are added for activation to obtain a microbead activation solution. The above bead activating solution was prepared in 16 parts, each part using beads of different codes.
3. Preparation of suspension of composite microbeads
Respectively adding 10-100ug of autoimmune liver disease antigen into 11 parts of microbead activating solution, mixing by vortex for 30 s, mixing by ultrasonic for 30 s, and shaking at 2-8 deg.C overnight. The above autoimmune liver disease antigens are as follows: SSA 52, anti-CENP B, AMA-M2, Actin, LKM-1, Sp100, gp210, Nup62, PML, LC-1, and SAL/LP. And preparing the detection micro-beads.
Taking 3 parts of microbead activating solution, respectively marking as a standard microbead 1, a standard microbead 2 and a standard microbead 3, adding 2-5ug/ml of human IgG into the calibration microbead 1, adding 6-10ug/ml of human IgG into the calibration microbead 2, adding 12-18ug/ml of human IgG into the calibration microbead 3, uniformly mixing by vortex for 30 seconds, uniformly mixing by ultrasonic for 30 seconds, and shaking at 2-8 ℃ overnight. Making into standard micro-beads.
Adding 8-15ug/ml of anti-human IgG into 1 part of the microbead activating solution, mixing uniformly by vortex for 30 seconds, mixing uniformly by ultrasonic waves for 30 seconds, and shaking uniformly at 2-8 ℃ overnight. Reference beads were made.
Taking 1 part of microbead activating solution, and adding no antigen as blank microbead.
And (3) carrying out suction filtration and washing on the microbeads, transferring the washed microbeads into a sample diluent, mixing, and finally fixing the volume to 120ml by using the sample diluent. And after the test is qualified, the mixture is subpackaged into 20 bottles, and each bottle contains 5.5 ml.
3. Fluorescent marker:
phycoerythrin-labeled goat anti-human IgG (IgG-PE): the initial concentration was 1 mg/ml.
Diluting with sample diluent to working concentration. The working concentration was 0.6 mg/L.
4. Concentrating the solution: the components and the formula are the same as the sample diluent, and the concentration is 10 times of the sample diluent. The preparation was 8L originally, and 800ml now.
5. Quality control product: positive quality control product: mixing the positive serums containing the multiple autoimmune liver disease antigens to obtain human mixed serums with positive analytes, and determining the concentration range of the positive analytes, wherein each batch of reagent has the specific concentration range. Negative quality control product: and mixing the negative serum to obtain a negative quality control product.
6. And respectively filling the prepared composite microbead suspension, fluorescent marker, sample diluent, washing solution and quality control product into reagent bottles, and then separating and packaging the reagent bottles collectively by using a kit.
Example 2:
the invention discloses a specific application of a multiple detection kit for an autoimmune liver antibody in simultaneous detection of multiple autoimmune liver disease antibodies in a human serum sample. The method comprises the following specific steps:
1. the components were removed from storage, protected from strong light, and allowed to return to room temperature (20-25 ℃).
2. And determining the total quantity of the quality control products and the samples required by the test. Negative controls were placed in well A1 and positive controls in well B1, each requiring a microwell (see sample alignment in Table 1).
a. The complex microbead suspension should be thoroughly mixed before use to obtain the best reading time. The most effective method is to shake the bead suspension with a vortex mixer for 30 seconds and then sonicate for 30 seconds.
b. The reagents should be thoroughly mixed when used. Suitable methods include placing the microplate on a vortexer and vortexing at about 800RPM for 30 seconds, or pipetting about 1/2 volumes of reaction liquid using a pipettor to repeat the pipetting at least 5 times.
TABLE 1
3. In dilution plates, the ratio of 1: the negative control, the positive control and each patient's serum sample were diluted at a dilution ratio of 21 (e.g., 10ul of serum sample +200ul of sample diluent). Note that: the correct operating requirements are: the sample and sample diluent are mixed uniformly as described above in section 2 b.
4. After determining the number of wells required for detection, 50ul of the complex bead suspension was added to each well of the filter plate using a multichannel pipettor or a reusable pipettor.
5. To each well of the filter plate was added 10ul of a 1: 21 diluted sample and quality control product. The correct operating requirements are: the material added should be mixed well with the suspension of composite microbeads in the wells as described above in section 2 b.
6. The filter plates were incubated at room temperature (20-25 ℃) for 30 minutes.
7. After incubation, the microbeads were washed with a vacuum pump as follows.
a. The filter plate was placed on the tray of the vacuum pump, the vacuum pump was turned on, the solution in the well was removed, leaving only the beads at the bottom of the plate.
b. The vacuum pump was turned off and 200ul of 1X wash solution was added to each well of the filter plate.
c. The solution was removed again using a vacuum pump.
d. The 7b, 7C steps were repeated, requiring a total of three washes.
8. After the last wash was complete, the bottom of the filter plate was gently blotted dry and the filter plate was allowed to air dry for 3-5 minutes before the next operation.
9. 150ul of fluorescent label was added to each well of the filter plate, and added to the filter plate sequentially according to the order and rate of sample addition. The correct operating requirements are: the added fluorescent label is mixed with the beads in the well uniformly, as described above in section 2 b. Alternatively, when the fluorescent label is added and mixed, the mixture may be transferred to an empty polystyrene reaction plate.
10. The filter plates were incubated at room temperature (20-25 ℃) for 30. + -.10 minutes.
11. A self-immune liver antibody template of a Luminex 200 multifunctional flow type dot matrix instrument of Luminex company in America is selected for analyzing test results. The results can be read from the filter plate or the reaction plate.
12. The filter plate reads within 60 minutes after the fluorescent marker incubation is complete. The filter plate was shaken for approximately 15 seconds before the reading. This optional step may reduce the time required to read the plate.
The multiple detection kit for the autoimmune liver antibody realizes the detection of multiple autoimmune liver disease antibodies on one human serum sample in the same hole at the same time.
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (7)
1. A kit for the multiple detection of autoimmune liver antibodies, the kit comprising: (1) a plurality of sealed reagent bottles respectively filled with the composite microbead suspension, the fluorescent marker, the sample diluent and the washing solution, and (2) a kit for separating and packaging the reagent bottles in a centralized manner; the microbeads are polystyrene microspheres with uniform size, and the polystyrene microspheres are dyed and numbered by different fluorescent dyes, so that microbeads with different codes are obtained; the composite microbead suspension comprises detection microbeads, standard microbeads, reference microbeads and blank microbeads, and specifically comprises the following components:
the detection beads are formed by respectively coating different autoimmune liver disease antigens on the surfaces of beads with different codes;
the standard microbeads comprise at least three microbeads with different concentration gradients of human IgG bound thereto;
the reference beads are anti-human IgG-bound beads;
the blank micro-bead is a micro-bead activating solution without any antigen added; the preparation method of the composite microbead suspension comprises the following steps: coating different autoimmune liver disease antigens with microbeads with different codes respectively, mixing to prepare detection microbeads, transferring the detection microbeads, standard microbeads, reference microbeads and blank microbeads to a sample diluent together, and mixing to obtain the composite microbead suspension; the fluorescent marker is phycoerythrin-labeled goat anti-human IgG;
the autoimmune liver disease antigens include: SSA 52, anti-CENP B, AMA-M2, Actin, LKM-1, Sp100, gp210, Nup62, PML, LC-1, and SAL/LP.
2. The multiple test kit for the detection of the autoimmune liver antibody according to claim 1, wherein the diameter of the microbeads is 5-7 microns.
3. The multiple detection kit for the autoimmune liver antibodies according to claim 1, wherein the composition and the content of each component in the sample diluent are as follows:
4. the multiple detection kit for the autoimmune liver antibodies according to claim 1, wherein the washing solution has the same composition as the sample dilution and has a concentration 8 to 10 times higher than the sample dilution.
5. The multiple detection kit for the autoimmune liver antibody according to claim 1, further comprising a reagent bottle containing quality control materials comprising a positive quality control material which is a human mixed serum containing an antibody corresponding to the antigen of autoimmune liver disease contained in the detection microbead and a negative quality control material which is a human negative serum negative for the antibody corresponding to the antigen of autoimmune liver disease.
6. The multiple detection kit for the autoimmune liver antibodies of claim 1, wherein the detection beads are prepared by the following method: preparing a microbead activating solution, adding any one autoimmune liver disease antigen according to the proportion that each milliliter of microbead contains 10-100ug of antigen, fully mixing, coating different autoimmune liver disease antigens, and using microbeads with different codes; the microbead activating solution is prepared by adding microbeads of 1000 unit volumes into a reaction tube, and adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride of 5-50 unit volumes and N-hydroxy thiosuccinimide of 5-50 unit volumes for activation.
7. The multiple detection kit for the autoimmune liver antibodies according to claim 1, wherein the standard microbeads are prepared by the following method: preparing the microbead activating solution, then adding human IgG according to the proportion that each milliliter of microbead contains 2-18ug of human IgG, and fully mixing; the reference microbead is prepared by the following method: preparing the microbead activating solution, then adding anti-human IgG into each milliliter of microbead according to the proportion that each milliliter of microbead contains 8-15ug of anti-human IgG, and fully mixing; the blank micro-bead is the micro-bead activating solution without any antigen; the microbead activating solution is prepared by adding microbeads of 1000 unit volumes into a reaction tube, and adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride of 5-50 unit volumes and N-hydroxy thiosuccinimide of 5-50 unit volumes for activation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810823210.9A CN109142743B (en) | 2018-07-25 | 2018-07-25 | Multiple detection kit for self-immune liver antibody |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810823210.9A CN109142743B (en) | 2018-07-25 | 2018-07-25 | Multiple detection kit for self-immune liver antibody |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109142743A CN109142743A (en) | 2019-01-04 |
| CN109142743B true CN109142743B (en) | 2022-02-08 |
Family
ID=64797805
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810823210.9A Active CN109142743B (en) | 2018-07-25 | 2018-07-25 | Multiple detection kit for self-immune liver antibody |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109142743B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113105545B (en) * | 2021-04-29 | 2022-04-08 | 四川携光生物技术有限公司 | II type self-immune liver LKM-1 and LC-1 composite quality control product and preparation method and application thereof |
| CN114113589A (en) * | 2021-10-25 | 2022-03-01 | 江苏纳迪芯生命科技研究院有限公司 | Chlamydia trachomatis and mycoplasma urealyticum antigen detection kit based on magnetic particle luminescence method |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1866013A (en) * | 2006-05-31 | 2006-11-22 | 山东省医药生物技术研究中心 | Liquid phase chip for parallel detection of autoantibodies, preparation method and application thereof |
| CN101226192A (en) * | 2008-02-03 | 2008-07-23 | 山东省医药生物技术研究中心 | Colour development protein chip using BCIP/NBT as substrate as well as application in detection of autoantibody thereof |
| CN101762688A (en) * | 2009-06-12 | 2010-06-30 | 华中科技大学同济医学院附属同济医院 | Experimental method for synchronously detecting relative content and absolute content of various antigen-specific associated antibodies |
| CN102707055A (en) * | 2012-06-11 | 2012-10-03 | 郑州安图绿科生物工程有限公司 | Kit for joint or single detection of autoimmune liver disease related antibody and detection method of kit |
| RU2477486C1 (en) * | 2011-11-23 | 2013-03-10 | Венера Эсанбаевна Сагынбаева | Diagnostic technique for crossed forms of autoimmune liver diseases |
| CN207396498U (en) * | 2017-10-24 | 2018-05-22 | 北京和杰创新生物医学科技有限公司 | Multi-series autoimmune disease antibody comprehensive detection film item |
Family Cites Families (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5380663A (en) * | 1984-12-24 | 1995-01-10 | Caribbean Microparticles Corporation | Automated system for performance analysis and fluorescence quantitation of samples |
| US4918004A (en) * | 1984-12-24 | 1990-04-17 | Caribbean Microparticles Corporation | Method of calibrating a flow cytometer or fluorescence microscope for quantitating binding antibodies on a selected sample, and microbead calibration kit therefor |
| CN101074950B (en) * | 2007-06-26 | 2011-02-09 | 东南大学 | Method and apparatus for inspecting gel chip |
| CN103983792B (en) * | 2014-05-29 | 2015-09-30 | 同昕生物技术（北京）有限公司 | A kind of immunoglobulin (Ig) lateral chromatography detection system |
| CN104459159A (en) * | 2014-12-23 | 2015-03-25 | 广州南杰生物技术有限公司 | Kit for detecting relevant autoantibody spectrum of autoimmune liver disease |
| CN107548461A (en) * | 2015-03-30 | 2018-01-05 | Hycor生物医学有限责任公司 | Immunoassay instrument system for the automation of the diagnostic measure that performs autoimmune disease and infectious diseases |
| WO2017138946A1 (en) * | 2016-02-11 | 2017-08-17 | Massachusetts Institute Of Technology | Multiplexed lateral flow assay |
-
2018
- 2018-07-25 CN CN201810823210.9A patent/CN109142743B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1866013A (en) * | 2006-05-31 | 2006-11-22 | 山东省医药生物技术研究中心 | Liquid phase chip for parallel detection of autoantibodies, preparation method and application thereof |
| CN101226192A (en) * | 2008-02-03 | 2008-07-23 | 山东省医药生物技术研究中心 | Colour development protein chip using BCIP/NBT as substrate as well as application in detection of autoantibody thereof |
| CN101762688A (en) * | 2009-06-12 | 2010-06-30 | 华中科技大学同济医学院附属同济医院 | Experimental method for synchronously detecting relative content and absolute content of various antigen-specific associated antibodies |
| RU2477486C1 (en) * | 2011-11-23 | 2013-03-10 | Венера Эсанбаевна Сагынбаева | Diagnostic technique for crossed forms of autoimmune liver diseases |
| CN102707055A (en) * | 2012-06-11 | 2012-10-03 | 郑州安图绿科生物工程有限公司 | Kit for joint or single detection of autoimmune liver disease related antibody and detection method of kit |
| CN207396498U (en) * | 2017-10-24 | 2018-05-22 | 北京和杰创新生物医学科技有限公司 | Multi-series autoimmune disease antibody comprehensive detection film item |
Non-Patent Citations (3)
| Title |
|---|
| Evaluation of a Multiplex Fluorescent Microsphere Immunoassay for the Determination of Epstein-Barr Virus Serologic Status;Thomas B. Martins等;《American Journal of Clinical Pathology》;20080131;第129卷(第1期);第34-41页 * |
| ZEUS AtheNA Multi-Lyte ANA-II Plus Test System;ZEUS Scientific, Inc.;《ZEUS AtheNA Multi-Lyte ANA-II Plus Test System》;20150915;https://www.zeusscientific.com/content/resources/A21101%2520AtheNA%2520ANA%2520II%2520Plus%2520English%2520Package%2520Insert.pdf * |
| 抗核抗体(Luminex微珠法)检测试剂盒;上海一滴准生物科技有限公司;《豆丁网》;20120921;http://www.docin.com/p-485734809.html * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN109142743A (en) | 2019-01-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US9488665B2 (en) | Magnetic particle tagged reagents and techniques | |
| EP3788372B1 (en) | Immunoassay for an automated system | |
| CN109725153B (en) | Homogeneous phase immunoassay method and application thereof | |
| JPH07508102A (en) | assay | |
| CN114107019B (en) | Microfluidic chip for simultaneously detecting nucleic acid and protein, detection method and application | |
| CN109142743B (en) | Multiple detection kit for self-immune liver antibody | |
| CN111521780B (en) | Kit for combined detection of hepatitis C virus antigen and antibody and application thereof | |
| CN111579781A (en) | Hepatitis C virus antibody detection kit, preparation method and detection method | |
| EP4182687A2 (en) | Immunoassay for sars-cov-2 neutralizing antibodies and materials therefor | |
| CN109633163B (en) | procalcitonin/C reactive protein two-in-one detection kit | |
| JP4975601B2 (en) | Reagent kit for HCV antibody measurement and HCV antibody measurement method | |
| KR20010025027A (en) | Immunoassay reagents and immunoassay method | |
| Al-Amri et al. | Qualitative and quantitative determination of MERS-CoV S1-specific antibodies using ELISA | |
| JPH09507577A (en) | Reaction column and method for multi-target simultaneous measurement | |
| CA2939789C (en) | Red blood cell detection | |
| CN111239403B (en) | Beta 2 microglobulin latex enhanced immunoturbidimetry kit and application | |
| CN108517316B (en) | Anti-human IgM monoclonal antibody, hybridoma cell strain and application thereof | |
| CN114258491A (en) | Immunoassay format for a fully novel total antibody assay | |
| CA2938935C (en) | Crossmatching blood samples | |
| JP2015184125A (en) | Immunoassay suppressing non-specific reaction | |
| WO2011022445A2 (en) | Improved multiplexed, microparticle-based assay for antibodies to antigens | |
| JPH0368346B2 (en) | ||
| JP4005093B2 (en) | Hepatitis C virus infection diagnostic agent and anti-HCV antibody detection method | |
| JP2008145181A (en) | Detecting method of parvovirus b19 | |
| JPH0772155A (en) | Method for measuring antigen-antibody reaction |
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 | ||
| CB02 | Change of applicant information |
Address after: 213149 No. 9 Changyang Road, West Taihu Science and Technology Industrial Park, Changzhou City, Jiangsu Province Applicant after: Zeus Life Technology (Changzhou) Co., Ltd. Address before: 213149 No. 9 Changyang Road, West Taihu Science and Technology Industrial Park, Changzhou City, Jiangsu Province Applicant before: One drop Biotechnology (Changzhou) Co., Ltd. |
|
| CB02 | Change of applicant information | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |