CN101566634A - Troponin I serum quick test kit (colloidal gold method) - Google Patents
Troponin I serum quick test kit (colloidal gold method) Download PDFInfo
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- CN101566634A CN101566634A CNA2008100364182A CN200810036418A CN101566634A CN 101566634 A CN101566634 A CN 101566634A CN A2008100364182 A CNA2008100364182 A CN A2008100364182A CN 200810036418 A CN200810036418 A CN 200810036418A CN 101566634 A CN101566634 A CN 101566634A
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Abstract
The invention belongs to the field of biotechnology and discloses a preparation method and a process of a troponin I serum quick test kit (colloidal gold method). A gold mark method is adopted to measure the content of cTnI in a human serum sample, the molecular weight of cardiac Troponin I (cTnI) is 22.5KD, and the cardiac Troponin I together with troponin T and troponin C form a structural compound which plays an important role in the transfer process of interaction of calcium signaling actin and myoglobulin between cells. The kit adopts an immunoluminometric assay; when a sample is put into a sample cell, the compound flows along a membrane and an antibody reacts in a detection zone by combining a binding pad and a gold antibody. If the sample has a marker with a certain concentration or higher concentration, a color strip appears in the detection zone. If the sample has no mark, the detection zone keeps colorless. The sample continues moving to a control area, a pink-rose color strip appears which proves that the test is normal and the result is valid. The invention has the advantages that the troponin I serum quick test kit has extremely stable performance and is convenient for users to accurately and quickly detect.
Description
Technical field
The invention provides a kind of Troponin I serum quick test kit and detection method of myocardial infarction fast detecting.
Background technology
Myocardial ischemic injury, particularly acute myocardial infarction (AMI) are one of principal diseases that threatens the human life, and clinical chemistry man and clinical science man strive to find always and explore good, the highly sensitive serum index of specificity.(cardiac troponin I cTnI) has myocardium specificity highly, when the cardiac muscle cell is impaired to cardiac muscle troponin I, cTnI time of occurrence morning in the blood, longer duration, closely related with myocardial damage degree and prognosis, therefore, it can be used as a kind of specific index of myocardial damage.
Summary of the invention
The invention provides a kind of Troponin I serum quick test kit and detection method of myocardial infarction fast detecting.
The present invention realizes by following technical solution: a kind of Troponin I serum quick test kit, it is characterized in that: a kind of Troponin I serum quick test kit, it is characterized in that: the molecular weight of myocardium calcium protein I (cTnI) is 22.5KD, and it constitutes structural composites together with TnT and TnC.Play an important role in the interactional conversion process of iuntercellular calcium signal actin-myosin in heart.People's cTnI and the form in the skeletal muscle are structurally different, many amino acid residues on its aminoterminal.Make cTnI become a specific mark.After myocardial infarction (AMI) took place, cTnI just was released in the blood circulation rapidly
1.cTnI the mark pre-service of monoclonal antibody
1.1 dialysis (purified water, 24 hours)
1.2 the centrifugal supernatant (4～10 ℃, 10000 rev/mins, 5 minutes) that goes
1.3 BCA rule concentration (material balance)
1.4 the adjusting (0.1mol/LK of PH
2CO
3Determining of amount)
1.5 determining of optimum mark amount
1.6 it is frozen that packing (5000 people's deals/prop up) ,-20 is spent
2. the preparation of reagent
2.1 the preparation of 1% trisodium citrate
2.2 the preparation of 1% gold chloride
2.3 the preparation of 0.01% gold chloride
2.4 the preparation of 0.1ml/L K2CO3
2.5 the preparation of 10% sodium chloride
2.6 the preparation of 1% Sodium azide
2.7 the preparation of 10% BSA
2.8 the preparation of 1mol/l PBS
2.9 the preparation of 0.05mol/l PBS
2.10 collaurum redissolve liquid preparation
3. the preparation of collaurum
3.1 measure 0.01% HAuCL4 solution
3.2 be heated to boiling
Add 1% citric acid three sodium solution rapidly, stir
(the HAuCL4 solution of ratio: 100ml 0.01% adds 1.6ml 1% citric acid three sodium solution)
3.3 continue heating 10 minutes
Be cooled to room temperature, return to original volume with ultrapure water
3.4 0.22 μ m filtering with microporous membrane degerming
3.5 inspect by ready samples, bottle, label
4. the preparation and the drying of cTnI immunity gold
4.1 the dilution of cTnI labelled antibody
Need 1 cTnI labelled antibody to calculate according to the wide cTnI test strips of every production 5000 person-portion 4mm, get the antibody of respective amount, be diluted to 0.5mg/ml with 0.05mol/l PBS.
4.2 lab scale before producing:
A gets the 1ml collaurum
B transfers PH (0.1mol/L K
2CO
3Amount, mixing 5min)
C adds CTnIAb1, mixing 10min according to producing instruction
D adds 100ul 10%BSA, mixing 10min
The centrifugal 1000 commentaries on classics/min of e, 30min (requiring environment temperature 4-10 ℃)
F30% redissolves, and immersion polyester film 0.8ml/ bar (0.6cm * 30cm)
20-30 ℃ of g temperature, dry 5 hours of relative humidity 30-35%
H send the check of quality inspection portion
4.3 cTnI antibody labeling
1) mark lab scale quality inspection quality inspection is qualified, gets collaurum, electromagnetic agitation
(need the 50ml collaurum to calculate to produce the wide Troponin I colloid gold test paper treaty of 1000 person-portion 5mm.)
2) transfer PH (0.1mol/L K
2CO
3Amount), mixing 5min
3) add CTnIAb1, mixing 10min according to the suitableeest labelled amount
4) add 100ul 10%BSA, mixing 10min
5) centrifugal 1000 commentaries on classics/min, 30min, 4-10 ℃
6) 30% redissolves, and soaks polyester film 25.3ml/ and opens (20cm * 30cm)
7) temperature 20-30 ℃, dry 5 hours of relative humidity 30-35%
8) take out a wide quality inspection portion that send of 0.6cm and do film gold matching detection
9) the product chamber was temporary in the middle of cTnI immunity gold sent
5.cTnI bag is by the preparation of plate
5.1 the dilution of coated antibody
Sheep anti mouse Ig0.05mol/LPBS is diluted to the 1.2mg/ml working concentration
The cTnI capture antibody is diluted to the 1.1mg/ml working concentration with 0.05mol/LPBS
5.2 bag is by preceding lab scale
1) respectively gets 20ul examination bag by the long film of 1 20cm, line concentration 1.0ul/cm
2) 2 ℃-8 ℃ are spent the night
3) pad pasting
4) dry 5 hours of temperature 20-30 ℃, relative humidity 30-35%
Send the check quality inspection of quality inspection portion qualified at last.
5.3 bag is produced
1) the cTnI bag is by the qualified list of lab scale quality inspection
2) sheep anti-mouse igg, cTnI capture antibody working concentration bag be by nature controlling line and detection line, line concentration 1.0ul/cm
3) 2 ℃-8 ℃ are spent the night
4) pad pasting
5) dry 5 hours of temperature 20-30 ℃, relative humidity 30-35%
6) taking out 1 plate send quality inspection portion to make film gold coupling
7) send middle product storehouse temporary after the sealing
The invention has the beneficial effects as follows the Troponin I serum quick test kit, performance is extremely stable, extremely convenient, the accurate fast detecting of energy that the user uses.
Description of drawings
Make the process chart of Troponin I serum quick test kit
Annotate: 1. the process of " ☆ " representative must be operated in the cleaning shop
2. the process of " ◆ " representative must be in temperature 20-30 degree, the operation of humidity 30-35 degree, and the cTnI bag also must be carried out at temperature 20-30 degree, humidity 30-35 degree by the dry run of the preparation of the preparation of the utmost point and cTnI immunity gold in addition.
3
Represent critical control point
Embodiment
The formulation operations of Troponin I serum quick test kit is as follows:
The molecular weight of myocardium calcium protein I (cTnI) is 22.5KD, and it constitutes structural composites together with TnT and TnC.Play an important role in the interactional conversion process of iuntercellular calcium signal actin-myosin in heart.People's cTnI and the form in the skeletal muscle are structurally different, many amino acid residues on its aminoterminal.Make cTnI become a specific mark.After myocardial infarction (AMI) took place, cTnI just was released in the blood circulation rapidly.The pattern that discharges identical with CK-MB (after AMI takes place 4-6 hour).Yet the level of CK-MB was recovered after 36-48 hour normally, and cTnI can continue to raise 6-10 days.In the normal health human body, the cTnI level is low-down, also detects less than cTnI in the impaired patient body of skeletal muscle.Therefore cTnI is a specific index of diagnosing myocardial infarction.
This kit adopts the double antibodies sandwich immunization, and after sample added sample cell, by pad and golden antibodies, compound reacted at surveyed area along membrane flow and antibody.Can detect concentration or denseer mark if having in the sample, will a colour band occur at detection zone.If do not have to keep colourless.Sample continues to move to the control area, pink-rosy colour band occurs, illustrates that the normal result of test is effective.
[name of product] Troponin I serum quick test kit (colloidal gold method)
[specification] 25 person-portions/box
[scope of application]
This kit is the single stage method external diagnosis reagent case on the immunochromatography basis.Myocardium calcium protein I when being used for to the generation myocardial infarction in the human serum carries out qualitative detection.
[detection principle] colloidal gold method
Use the double antibody sandwich method principle, with collaurum as cue mark, crosslinked anti-troponin I antibody, another antibody is capture antibody, combine with Troponin I in the blood and develops the color.During test, sample moves to detection zone and Quality Control district, if contain Troponin I in the sample, it will form antigen antibody complex with the golden labeling antibody in the reagent, again with detection zone in corresponding capture antibody form mauve colour band.If the Troponin I level is lower than lowest detectable limit in the sample, just can not occur by color band in the detection zone.Under any circumstance, Quality Control district colour band all should occur.The appearance of Quality Control district colour band shows that reactive system is effective.
[main constituent]
Test card and a drying agent, a dropper are loaded in the aluminium foil bag together, and each kit has 25 identical aluminium foil bags.
[providing material for oneself]
1, serum collecting device
2, time set
[sample requirement]
1, must under the standard test room environmental, gather serum specimen.
2, preferably just do detection after the collection of specimens at once.As not detecting in 24 hours, serum specimen should be freezing, made sample be returned to room temperature before test.
3,0.1% Sodium azide can be added as antiseptic, test findings can be do not influenced.
[quality control method]
1. control line is internal reference reagent and step control.As long as the effective control line of proper operation reagent always occurs.
2. suggestion all uses reference substance to guarantee the validity of kit at every turn.Reference substance, buyable are not provided in this kit.
[operation steps]
1. before using, all reagent composition and patient's sample are placed room temperature.
2. test card is taken out from aluminium foil bag.
3. draw 150 μ l samples with micropipettor.
4. pipettor is vertical, adds sample 2-3 and drips (100-150 μ l) to sample cell.
5.15 minute reading displayed result.
[to the explanation of test findings]
Positive:
Occur two colour bands after 15 minutes, illustrate that positive test is effective as a result.Promptly can read the result as long as have colour band at surveyed area.
Annotate: very two bands can appear in the cTnI of low concentration when being longer than 15 minutes.
Negative:
If do not have band and there is band the control area, the negative and test of result is described effectively at surveyed area.
Invalid test:
If without any band, invalidate the test is described in the control area, sample is used a new card and is tested again.
[product performance index]
Positive findings appears in this kit when cTnI 〉=1.5ng/ml.
[points for attention]
1. when handling sample, please wear disposable glove.
2. when testing, test card is answered horizontal positioned, tilts.
Application of sample after 15 minutes result displayed do not have diagnostic significance.
4. end of test (EOT) is please thoroughly cleaned both hands residue sample and discarded object and is used 84 medicining liquid dippings of dilution in 1: 25 and just can abandon more than 30 minutes.
5. a suction pipe can only be drawn a sample, in order to avoid cross pollution.
6. if drip sample, should disposablely rapidly join in the sample cell, avoid interrupted adding with the micropipet head.
7. this reagent result only supplies auxiliary diagnosis qualitatively, and experimental result is explained to be needed to combine with clinical symptoms.
[storage] 2-8 ℃ of preservation.
[term of validity] 12 months.
[medicine equipment certificate of registry numbering]
[product standard numbering] Q/YZ 021-2006
Claims (4)
1. the molecular weight of myocardium calcium protein I (cTnI) is 22.5KD, it constitutes structural composites together with TnT and TnC, play an important role in the interactional conversion process of iuntercellular calcium signal actin-myosin in heart, this kit adopts the double antibodies sandwich immunization, after sample adds sample cell, by pad and golden antibodies, compound reacts at surveyed area along membrane flow and antibody.Can detect concentration or denseer mark if having in the sample, will a colour band occur at detection zone.If do not have to keep colourless.Sample continues to move to the control area, pink-rosy colour band occurs, illustrates that the normal result of test is effective.
2. Troponin I serum quick test kit according to claim 1 (colloidal gold method) is characterized in that: the molecular weight of myocardium calcium protein I (cTnI) is 22.5KD, and it constitutes structural composites together with TnT and TnC.
3. the preparation of Troponin I serum quick test kit according to claim 1 (colloidal gold method), it is characterized in that: 1% trisodium citrate, 1% gold chloride, 0.01% gold chloride, 0.1ml/LK2CO3,10% sodium chloride, 1% Sodium azide, 10% BSA, 1mol/l PBS, 0.05mol/lPBS, collaurum redissolution liquid.
4. Troponin I serum quick test kit according to claim 1 (colloidal gold method) comprises following processing step:
The pre-service of monoclonal antibody of first step cTnI mark
1.1 dialysis (purified water, 24 hours)
1.2 the centrifugal supernatant (4 ~ 10 ℃, 10000 rev/mins, 5 minutes) that goes
1.3BCA rule concentration (material balance)
1.4PH adjusting (0.1mol/LK2CO3 amount determine)
1.5 determining of optimum mark amount
1.6 it is frozen that packing (5000 people's deals/prop up) ,-20 is spent
The preparation of the second step collaurum
2.1 measure 0.01% HAuCL4 solution (being heated to boiling)
2.2 add 1% citric acid three sodium solution rapidly, stir
(the HAuCL4 solution of ratio: 100ml 0.01% adds 1.6ml 1% citric acid three sodium solution) (continuing heating 10 minutes)
2.3 be cooled to room temperature, return to original volume with ultrapure water
2.40.22 μ m filtering with microporous membrane degerming
2.5 inspect by ready samples, bottle, label
The preparation and the drying of the 3rd step cTnI immunity gold
3.1cTnI the dilution of labelled antibody
Need 1 cTnI labelled antibody to calculate according to the wide cTnI test strips of every production 5000 person-portion 4mm, get the antibody of respective amount, be diluted to 0.5mg/ml with 0.05mol/l PBS.
3.2 lab scale before producing:
Get the 1ml collaurum
1) transfers PH (amount of 0.1mol/L K2CO3, mixing 5min)
2) add CTnIAb1, mixing 10min according to producing instruction
3) add 100ul 10%BSA, mixing 10min
4) centrifugal 1000 commentaries on classics/min, 30min (requiring environment temperature 4-10 ℃)
5) 30% redissolves, and immersion polyester film 0.8ml/ bar (0.6cm * 30cm)
6) temperature 20-30 ℃, dry 5 hours of relative humidity 30-35%
7) send the check of quality inspection portion
3.3cTnI antibody labeling
1) mark lab scale quality inspection quality inspection is qualified
2) get collaurum, electromagnetic agitation
(need the 50ml collaurum to calculate to produce the wide Troponin I colloid gold test paper treaty of 1000 person-portion 5mm.)
3) transfer PH (amount of 0.1mol/L K2CO3), mixing 5min
4) add CTnIAb1, mixing 10min according to the suitableeest labelled amount
5) add 100ul 10%BSA, mixing 10min
6) centrifugal 1000 commentaries on classics/min, 30min, 4-10 ℃
7) 30% redissolves, and soaks polyester film 25.3ml/ and opens (20cm * 30cm)
8) temperature 20-30 ℃, dry 5 hours of relative humidity 30-35%
9) take out a wide quality inspection portion that send of 0.6cm and do film gold matching detection
10) the product chamber was temporary in the middle of cTnI immunity gold sent
The 4th step cTnI bag is by the preparation of plate
4.1 the dilution of coated antibody
Sheep anti mouse Ig0.05mol/LPBS is diluted to the 1.2mg/ml working concentration
The cTnI capture antibody is diluted to the 1.1mg/ml working concentration with 0.05mol/LPBS
4.2 bag is by preceding lab scale
1) respectively gets 20ul examination bag by the long film of 1 20cm, line concentration 1.0ul/cm
2) 2 ℃-8 ℃ are spent the night
3) pad pasting
4) dry 5 hours of temperature 20-30 ℃, relative humidity 30-35% send the check quality inspection of quality inspection portion qualified.
4.3 bag is produced
1) the cTnI bag is by the qualified list of lab scale quality inspection
2) sheep anti-mouse igg, cTnI capture antibody working concentration bag be by nature controlling line and detection line, line concentration 1.0ul/cm
3) 2 ℃-8 ℃ are spent the night
4) pad pasting
5) dry 5 hours of temperature 20-30 ℃, relative humidity 30-35%
6) taking out 1 plate send quality inspection portion to make film gold coupling
7) send middle product storehouse temporary after the sealing
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008100364182A CN101566634A (en) | 2008-04-22 | 2008-04-22 | Troponin I serum quick test kit (colloidal gold method) |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008100364182A CN101566634A (en) | 2008-04-22 | 2008-04-22 | Troponin I serum quick test kit (colloidal gold method) |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN101566634A true CN101566634A (en) | 2009-10-28 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNA2008100364182A Pending CN101566634A (en) | 2008-04-22 | 2008-04-22 | Troponin I serum quick test kit (colloidal gold method) |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102183633A (en) * | 2011-02-24 | 2011-09-14 | 南京基蛋生物科技有限公司 | Colloidal gold labeling method |
| CN102192986A (en) * | 2010-03-19 | 2011-09-21 | 胡卫红 | Cardiac troponin I detection reagent and preparation method thereof and application thereof |
| CN102353787A (en) * | 2011-09-27 | 2012-02-15 | 山东莱博生物科技有限公司 | Colloidal gold test paper strip for semiquantitatively detecting concentration of GFAP (Glial Fibrillary Acidic Protein) in human serum |
| CN102426246A (en) * | 2011-08-31 | 2012-04-25 | 内蒙古科慧生物科技有限责任公司 | Human troponin I quantitative determination kit and detection method |
| CN102539787A (en) * | 2012-01-10 | 2012-07-04 | 上海一滴准生物科技有限公司 | Rapid troponin I serum detection reagent kit (colloidal golden method) |
| CN103380377A (en) * | 2011-02-25 | 2013-10-30 | 三菱化学美迪恩斯株式会社 | Method for measuring myocardial troponin |
| CN103913580A (en) * | 2014-04-23 | 2014-07-09 | 齐鲁工业大学 | Nanotechnology for myocardial infarction diagnosis and device adopting same |
| CN103930783A (en) * | 2011-11-16 | 2014-07-16 | 皇家飞利浦有限公司 | Particle repulsion to enhance surface contact in magnetic particle immunoassays |
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Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102192986A (en) * | 2010-03-19 | 2011-09-21 | 胡卫红 | Cardiac troponin I detection reagent and preparation method thereof and application thereof |
| CN102192986B (en) * | 2010-03-19 | 2014-06-11 | 成都盛泰尔生物医药科技有限公司 | Cardiac troponin I detection reagent and preparation method thereof and application thereof |
| CN102183633A (en) * | 2011-02-24 | 2011-09-14 | 南京基蛋生物科技有限公司 | Colloidal gold labeling method |
| CN102183633B (en) * | 2011-02-24 | 2013-05-15 | 南京基蛋生物科技有限公司 | Colloidal gold labeling method |
| CN103380377A (en) * | 2011-02-25 | 2013-10-30 | 三菱化学美迪恩斯株式会社 | Method for measuring myocardial troponin |
| US10634681B2 (en) | 2011-02-25 | 2020-04-28 | Lsi Medience Corporation | Method for measuring cardiac troponin |
| CN103380377B (en) * | 2011-02-25 | 2016-01-27 | 美迪恩斯生命科技株式会社 | The assay method of cardiac troponin |
| CN102426246A (en) * | 2011-08-31 | 2012-04-25 | 内蒙古科慧生物科技有限责任公司 | Human troponin I quantitative determination kit and detection method |
| CN102353787B (en) * | 2011-09-27 | 2013-03-06 | 山东莱博生物科技有限公司 | Colloidal gold test paper strip for semiquantitatively detecting concentration of GFAP (Glial Fibrillary Acidic Protein) in human serum |
| CN102353787A (en) * | 2011-09-27 | 2012-02-15 | 山东莱博生物科技有限公司 | Colloidal gold test paper strip for semiquantitatively detecting concentration of GFAP (Glial Fibrillary Acidic Protein) in human serum |
| CN103930783A (en) * | 2011-11-16 | 2014-07-16 | 皇家飞利浦有限公司 | Particle repulsion to enhance surface contact in magnetic particle immunoassays |
| CN102539787A (en) * | 2012-01-10 | 2012-07-04 | 上海一滴准生物科技有限公司 | Rapid troponin I serum detection reagent kit (colloidal golden method) |
| CN103913580A (en) * | 2014-04-23 | 2014-07-09 | 齐鲁工业大学 | Nanotechnology for myocardial infarction diagnosis and device adopting same |
| CN109557313A (en) * | 2019-01-04 | 2019-04-02 | 上海黄锦科技有限公司 | A kind of household HPV detection device and its detection method |
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Open date: 20091028 |