CN109307765A - Type pepsinogen II detection kit - Google Patents
Type pepsinogen II detection kit Download PDFInfo
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- CN109307765A CN109307765A CN201811446772.2A CN201811446772A CN109307765A CN 109307765 A CN109307765 A CN 109307765A CN 201811446772 A CN201811446772 A CN 201811446772A CN 109307765 A CN109307765 A CN 109307765A
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- pepsinogen
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- detection kit
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/76—Chemiluminescence; Bioluminescence
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/573—Immunoassay; Biospecific binding assay; Materials therefor for enzymes or isoenzymes
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
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Abstract
The invention discloses a kind of type pepsinogen II detection kits, magnetic particle including being coated with Pepsinogen II monoclonal antibody, enzyme dilution containing horseradish peroxidase (HRP) label Pepsinogen II monoclonal antibody, calibration object and sample diluting liquid containing Pepsinogen II antigen；Contain blocking agent ingredient in the sample diluting liquid；The blocking agent is the human anti-mouse antibody of one or more neutralization HAMA interference.The advantage of the invention is that highly sensitive chemiluminescence and magnetic particle technology of preparing are combined, the full-automatic detection analysis instrument of AutoLumo is matched, it is full-automatic to realize detection；It is added to blocking agent ingredient in sample diluting liquid simultaneously, heterophil antibody in sample can be eliminated, substantially increase the anti-interference ability of kit.
Description
Technical field
The present invention relates to Measurements for Biotechnique, more particularly, to a kind of type pepsinogen II detection kit.
Background technique
Propepsin (PG) is the inactive precursor of pepsin in gastric juice (proteolytic enzyme), for by 375 amino
The polypeptide chain of acid composition, average molecular mass are 42000 Da, have 7 groups of pepsin isodynamic enzymes in people's stomach lining
It is former.Pepsinogen Cgene (PG I), pepsin can be divided into according to distribution in biochemical property, immunogenicity, cell origin and tissue
Former II (PG II) two subgroups, 1-5 component immunogenicity is approximate, and referred to as PG I is mainly thin by the mucus neck of the chief cell of fundus gland
Intracrine；Component 6-7 is referred to as PG II, except by body of stomach and gastric mucosa secrete the cell secretes of acid gland in addition to, secrete the glutinous of acid gland
The mucilage cell of the pyloric gland of liquid neck cell, cardiac gland and antrum and the Brunner gland of duodenum upper section can also generate PG
II, the PG II of stomach lining synthesis is about the 25% of total amount.Stomach is almost the sole source of PG, and the PG after synthesis largely enters stomach
Chamber, activation enters blood through stomach lining capillary at propepsin, only a small amount of PG (about 1%) under acidic gastric juice effect
Liquid circulation, therefore serum PG I and PG II reflect the secreting function of stomach lining different parts.When pathological change occurs for stomach lining,
Serum PG content also changes therewith.Gastric juice and serum PG level and living tissue pathological change result are often consistent, therefore, monitoring
The concentration of PG can be used as the means of monitoring stomach lining state in serum.During fundus gland mucosal atrophy, the master of PG I is secreted
Leukopenia, pyloric gland cytosis, to cause the reduction of the ratio of PG I and PG II.Therefore the ratio of PG I and PG II can be made
For the indication of fundus gland mucosal atrophy, immunological method detection PG I and PG II is horizontal and the ratio of PG I and PG II is combined to can be used for
Screening fundus gland mucosal atrophy disease, such as gastritis, gastric cancer are related.
In the prior art, the detection method of PG II includes enzyme linked immunosorbent assay (ELISA) (ELISA), immunoturbidimetry, chemistry
Luminescent immunoassay (CLIA) etc..Enzyme-linked immunosorbent assay for measuring operating process is complicated, and minute is longer, influence factor
It is more, and be not suitable for the detection of emergency treatment sample；Immunoturbidimetry measurement when antigen or excessive amount of antibody, may occur in which soluble compound
Object causes error, influences vulnerable to piarhemia.Though chemiluminescence immunoassay has the multiple advantages such as the reaction time is short, resist dry
It is lower the performances such as to disturb, this is because most of antibody used in immunity detection reagent derives from experimental animal, the thermophilic opposite sex at present
With Fc the and Fab epitope of the immunoglobulin of many animals non-specific binding can occur for antibody, so that interference detection results go out
Existing false sun or missing inspection.
Summary of the invention
The purpose of the present invention is to provide a kind of type pepsinogen II detection kit, it is greatly improved its anti-interference energy
Power.
To achieve the above object, the present invention can take following technical proposals:
Type pepsinogen II detection kit of the present invention, the magnetic including being coated with Pepsinogen II monoclonal antibody are micro-
Grain, the enzyme dilution containing horseradish peroxidase (HRP) label Pepsinogen II monoclonal antibody, contains propepsin
The calibration object and sample diluting liquid of II antigen；Contain blocking agent ingredient in the sample diluting liquid；The blocking agent be it is a kind of or
A variety of human anti-mouse antibody for neutralizing HAMA interference.
The formula composition of the sample diluting liquid is 0.01 ~ 0.2 mol/L of buffer, 0.1% ~ 0.5%(w/v of preservative),
1% ~ 10%(w/v of protected protein), 1% ~ 10%(w/v of EDTA), blocking agent 200 ~ 500 ng/mL, pigment 0.001%(g/v).
The EDTA is disodium ethylene diamine tetraacetate or/and EDTAP dipotassium ethylene diamine tetraacetate.
It is coated with the magnetic particle of Pepsinogen II monoclonal antibody in the present invention, contains horseradish peroxidase-labeled stomach
The enzyme dilution of II monoclonal antibody of proproteinase, four groups of the calibration object containing Pepsinogen II antigen and sample diluting liquid
Dilution used in point is any one of PBS, MES, MOPS, pH=6.0~8.0.
Protected protein in the dilution is one or both of bovine serum albumin(BSA), cow's serum, casein.It is described
Preservative in dilution is P300, Bro, MIT, NaN3Any combination.
The magnetic particle is that Surface mulch has high molecular material Fe3O4Or Fe2O3Magnetic particle, and magnetic particle surface contains
There are one or more activity functional groups.
The advantage of the invention is that highly sensitive chemiluminescence and magnetic particle technology of preparing are combined, it is mating to make
With the full-automatic detection analysis instrument of AutoLumo, it is full-automatic to realize detection；Blocking agent is added in sample diluting liquid simultaneously
Ingredient can eliminate heterophil antibody in sample, substantially increase the anti-interference ability of kit, the principle is as follows:
EDTA is also known as ethylenediamine tetra-acetic acid, is commonly sodium salt or sylvite, can eliminate in fresh sample in immune detection
Complement, to avoid new and old differences between samples；Complement causes the reason of new and old differences between samples to be to participate in classical complement activation system
System, complement system are known as the activation of complement system from proenzyme condition conversion at the process with enzymatic activity state, and wherein one
Approach by antigen-antibody complex combination C1q starting complement activation is the Classical pathway of complement system.Antigen and anti-
After body combines, C1q can identify the complement combining site on antibody, and be combined with the Fc of antibody section, to generate to testing result dry
It disturbs, as shown in Figure 1.
Detailed description of the invention
Fig. 1 is detection principle diagram of the invention.
Fig. 2 is that kit of the present invention is compared with contrast agents box coincidence rate.
Specific embodiment
More detailed explanation is done to the present invention below by specific embodiment, in order to the reason of those skilled in the art
Solution.
Embodiment 1 prepares type pepsinogen II detection kit
1, magnetic particle suspension is prepared
After 1.08 μm of partial size of magnetic particle stoste is mixed well, washed, glutaraldehyde activated dose of 10 ~ 50 mg/ml is added, mixes
It is even to shake, after washing, the pepsinogen Cgene antibody of 0.1 ~ 0.5 μ g/ person-portion is added, mixes concussion, finally uses and contains PBS and 1%
The closing of ~ 5% bovine serum albumin(BSA) confining liquid, 2-8 DEG C of preservation.
2, enzyme dilution is prepared
1% ~ 5% bovine serum albumin(BSA) is added into prepared MES buffer, is enzyme dilution after mixing, HRP is marked
Antibody according to 1:500 ~ 1:5000 ratio be added mix, 2 ~ 8 DEG C preservation.
3, calibration object is prepared
1% ~ 5% bovine serum albumin(BSA) is added into prepared MOPS buffer, being uniformly mixed is calibration object dilution, uses school
Quasi- product dilution by II antigen diluent of PG at 6 gradients, 0ng/ml, 10ng/ml, 25ng/ml, 50 ng/ml, 100 ng/ml,
200 ng/ml are distributed into 1 ml/ bottles, 2-8 DEG C of preservation.
4, sample diluting liquid is prepared
Its component see the table below 1:
The examination of 2 Sample preservation condition of embodiment
Pepsinogen II gradient fresh sample 10 is taken, is detected respectively in 0h, 8h, 2d, 3d, 5d, each time detection knot
Fruit is compared with 0h result, examines Almost Sure Sample Stability.Sample is saved at 2 ~ 8 DEG C during experiment.As a result such as the following table 2:
The kit Sample preservation time study of the present invention of table 2
Table 2 the results show that kit of the present invention the Sample preservation time up to 5 days of 2~8 DEG C.
The performance evaluation of the kit of the present invention of embodiment 3
1, sensitivity technique
LOB prepares 5 parts of clinical samples close to 0 value, and each sample is repeated 3 times, does in total 4 days, obtains 60 data；LOD,
Prepare the range of clinical sample that 5 parts of concentration ranges are 1-4 times of LOB, each sample is repeated 3 times, does in total 4 days, obtains 60 numbers
According to；FS: using LOD test in data, 5 concentration samples survey 3 times daily, survey 4 days in total, and each sample obtains 12 knots
Fruit calculates mean value, SD and the CV% of each sample, and the concentration closest to 20% is Functional Sensitivity；The testing result of two batches is such as
Shown in table 3.
The kit sensitivity technique of the present invention of table 3
As can be seen from Table 3, first concentration that can accurately detect is 0.85 ng/mL, and second batch can be examined accurately
Concentration out is 0.92 ng/mL, is all satisfied requirement.
2, precision detects
Two batches reagent is taken to carry out Precision Experiment, respectively with the high/medium/low value sample of mating quality-control product and clinical high/medium/low value sample
This, each to survey 20 variations for calculating measurement concentration, measurement result is as shown in table 4, table 5.
The detection of the kit accuracy of the present invention of table 4
The detection of the kit accuracy of the present invention of table 5
Table 4,5 data result of table show: the coefficient of variation of kit of the present invention is respectively less than 5%.
3, accuracy determination
Since at present there is no countries or international standard substance for accuracy, herein using the accurate of the method kits for evaluation of recovery test
Degree.
10 parts of serum high level in the linear range are mixed with the pooled serum sample close to 0 value according to the ratio of 1:9,
Seek the rate of recovery.
The kit accuracy of the present invention of table 6 detection
As can be seen from Table 6, the rate of recovery of kit of the present invention is all satisfied requirement between 85%~115%.
4, clinical performance is examined
96 clinical diagnosis disease of stomach samples are measured simultaneously with kit of the present invention and external Abbott Laboratories' kit, include chronic stomach
The ratio result of inflammation, acute gastritis, gastric ulcer etc., joint PG I, PG II and PG I and PG II carry out the analysis of yin-yang coincidence rate, as a result
As shown in table 7.
The kit of the present invention of table 7 is compared with contrast agents box coincidence rate
It can be seen that kit of the present invention from 7 measurement result of table and Abbott Laboratories' positive coincidence rate reach 94.7%, negative match-rate reaches
To 100%, total coincidence rate reaches 98.9%；PG II is shown in attached drawing 2:y=0.955x+0.0354, r with Abbott Laboratories' correlation2=0.9937, it says
Bright kit can be used to detect disease of stomach, and carry out risk assessment according to testing result, can be under patient one
Step treatment offer scheme, reduces the incidence of early carcinoma of stomach.
Claims (7)
1. a kind of type pepsinogen II detection kit, the magnetic particle including being coated with Pepsinogen II monoclonal antibody, contain
There is the enzyme dilution of horseradish peroxidase-labeled Pepsinogen II monoclonal antibody, the school containing Pepsinogen II antigen
Quasi- product and sample diluting liquid；It is characterized by: containing blocking agent ingredient in the sample diluting liquid；The blocking agent be it is a kind of or
A variety of human anti-mouse antibody for neutralizing HAMA interference.
2. type pepsinogen II detection kit according to claim 1, it is characterised in that: the sample diluting liquid is matched
Side's group becomes 0.01 ~ 0.2 mol/L of buffer, preservative 0.1% ~ 0.5%, protected protein 1% ~ 10%, EDTA 1% ~ 10%, blocking
200 ~ 500 ng/mL of agent, pigment 0.001%.
3. type pepsinogen II detection kit according to claim 2, it is characterised in that: the EDTA is ethylenediamine tetraacetic
Acetic acid disodium or/and EDTAP dipotassium ethylene diamine tetraacetate.
4. type pepsinogen II detection kit according to claim 1, it is characterised in that: described to be coated with pepsin
The magnetic particle of former II monoclonal antibody, the enzyme dilution containing horseradish peroxidase-labeled Pepsinogen II monoclonal antibody
Liquid, dilution used in four components of the calibration object containing Pepsinogen II antigen and sample diluting liquid be PBS, MES,
Any one of MOPS, pH=6.0~8.0.
5. type pepsinogen II detection kit according to claim 4, it is characterised in that: the protection in the dilution
Albumen is one or both of bovine serum albumin(BSA), cow's serum, casein.
6. type pepsinogen II detection kit according to claim 4, it is characterised in that: the anti-corrosion in the dilution
Agent is P300, Bro, MIT, NaN3Any combination.
7. type pepsinogen II detection kit according to claim 1, it is characterised in that: the magnetic particle covers for surface layer
It is stamped high molecular material Fe3O4Or Fe2O3Magnetic particle, and one or more activity functional groups are contained on magnetic particle surface.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811446772.2A CN109307765A (en) | 2018-11-29 | 2018-11-29 | Type pepsinogen II detection kit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811446772.2A CN109307765A (en) | 2018-11-29 | 2018-11-29 | Type pepsinogen II detection kit |
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| Publication Number | Publication Date |
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| CN109307765A true CN109307765A (en) | 2019-02-05 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201811446772.2A Withdrawn CN109307765A (en) | 2018-11-29 | 2018-11-29 | Type pepsinogen II detection kit |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110501493A (en) * | 2019-08-28 | 2019-11-26 | 郑州安图生物工程股份有限公司 | A kind of detection kit of hemadsorption virus type 1's IgM antibody |
| CN110702906A (en) * | 2019-10-11 | 2020-01-17 | 郑州安图生物工程股份有限公司 | Kit for detecting anti-mutant citrullinated vimentin antibody |
| CN110824175A (en) * | 2019-11-28 | 2020-02-21 | 郑州安图生物工程股份有限公司 | Sample dilution composition for detection of saccharide antigen 72-4, detection reagent and kit comprising detection reagent |
| CN111781386A (en) * | 2020-08-21 | 2020-10-16 | 武汉生之源生物科技股份有限公司 | Antibody-coated magnetic particle buffer solution, gastrin 17 determination kit and preparation method thereof |
| CN113495156A (en) * | 2020-03-20 | 2021-10-12 | 郑州达诺生物技术有限公司 | Enzyme conjugate diluent, AMH quantitative detection kit and use method thereof |
| CN117825708A (en) * | 2024-03-04 | 2024-04-05 | 宁波美康盛德生物科技有限公司 | sFlt-1 detection kit |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104914092A (en) * | 2015-05-22 | 2015-09-16 | 必欧瀚生物技术（合肥）有限公司 | Pepsinogen II enzymatic chemiluminescence immunoassay kit |
| CN105548565A (en) * | 2015-12-30 | 2016-05-04 | 深圳市新产业生物医学工程股份有限公司 | Kit for detecting trypanosoma cruzi antibody as well as preparation and application thereof |
-
2018
- 2018-11-29 CN CN201811446772.2A patent/CN109307765A/en not_active Withdrawn
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104914092A (en) * | 2015-05-22 | 2015-09-16 | 必欧瀚生物技术（合肥）有限公司 | Pepsinogen II enzymatic chemiluminescence immunoassay kit |
| CN105548565A (en) * | 2015-12-30 | 2016-05-04 | 深圳市新产业生物医学工程股份有限公司 | Kit for detecting trypanosoma cruzi antibody as well as preparation and application thereof |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110501493A (en) * | 2019-08-28 | 2019-11-26 | 郑州安图生物工程股份有限公司 | A kind of detection kit of hemadsorption virus type 1's IgM antibody |
| CN110702906A (en) * | 2019-10-11 | 2020-01-17 | 郑州安图生物工程股份有限公司 | Kit for detecting anti-mutant citrullinated vimentin antibody |
| CN110702906B (en) * | 2019-10-11 | 2022-03-01 | 郑州安图生物工程股份有限公司 | Kit for detecting anti-mutant citrullinated vimentin antibody |
| CN110824175A (en) * | 2019-11-28 | 2020-02-21 | 郑州安图生物工程股份有限公司 | Sample dilution composition for detection of saccharide antigen 72-4, detection reagent and kit comprising detection reagent |
| CN113495156A (en) * | 2020-03-20 | 2021-10-12 | 郑州达诺生物技术有限公司 | Enzyme conjugate diluent, AMH quantitative detection kit and use method thereof |
| CN111781386A (en) * | 2020-08-21 | 2020-10-16 | 武汉生之源生物科技股份有限公司 | Antibody-coated magnetic particle buffer solution, gastrin 17 determination kit and preparation method thereof |
| CN117825708A (en) * | 2024-03-04 | 2024-04-05 | 宁波美康盛德生物科技有限公司 | sFlt-1 detection kit |
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Application publication date: 20190205 |