CN106093426A - A kind of test kit measuring bladder chalone C and preparation method thereof - Google Patents
A kind of test kit measuring bladder chalone C and preparation method thereof Download PDFInfo
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- CN106093426A CN106093426A CN201610376097.5A CN201610376097A CN106093426A CN 106093426 A CN106093426 A CN 106093426A CN 201610376097 A CN201610376097 A CN 201610376097A CN 106093426 A CN106093426 A CN 106093426A
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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
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
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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
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/34—Genitourinary disorders
- G01N2800/347—Renal failures; Glomerular diseases; Tubulointerstitial diseases, e.g. nephritic syndrome, glomerulonephritis; Renovascular diseases, e.g. renal artery occlusion, nephropathy
Abstract
The invention discloses a kind of test kit measuring bladder chalone C and preparation method thereof, including reagent R1 independent of each other and reagent R2 biliquid component composition test kit: reagent R1: buffer, inorganic ion, accelerator, stabilizer, preservative, reagent R2: buffer, stabilizer, suspending agent, surfactant, polyoxyethylene polyoxypropylene copolymer, preservative, latex are coated anti-human bladder chalone C antibody, and preparation method is: prepare reagent according to constituent content；Sample to be tested is mixed with reagent R1 and reagent R2 so that it is fully react；Reacted absorbance difference is measured with automatic clinical chemistry analyzer；The concentration of bladder chalone C in sample is calculated according to absorbance changing value.The present invention has accuracy advantages of higher.
Description
Technical field
The present invention relates to medical science and technological field of biochemistry, a kind of test kit measuring bladder chalone C and
Its preparation method.
Background technology
Bladder chalone C (Cys-C), be a kind of cystatin, also referred to as γ-trace albumin and γ-after
Globulin, is widely present in nucleated cell and the body fluid of various tissue, is a kind of low-molecular-weight, alkalescence non-glycated protein,
Molecular weight is 13.3KD, is made up of 122 amino acid residues, can be produced by all nucleated cell of body, and generation rate is constant, circulation
In bladder chalone C be only eliminated through glomerular filtration, be a kind of endogenous mark reflecting that glomerular filtration rate changes, and
Heavily absorbing at proximal convoluted tubule, but by complete metabolic breakdown after heavily absorbing, do not return blood, therefore, its blood level is by glomerule
Filter and determine, and be independent of any foeign element, such as sex, age, the impact of diet, be that a kind of reflection glomerular filtration rate becomes
The preferable homology mark changed.
Under normal circumstances, the bladder chalone C (Cys-C) concentration in serum and blood plasma is 0.51-1.09mg/L, works as renal function
Time impaired, bladder chalone C (Cys-C) concentration in blood changes with glomerular filtration rate change, and during renal failure, glomerule is filtered
Rate of crossing declines, and Cys-C concentration in blood can increase more than 10 times；If glomerular filtration rate is normal, and when renal tubular function is not normal,
Cys-C can be hindered to absorb at renal tubules and decompose rapidly, make the concentration in urine increase more than 100 times.
The main method of Clinical detection bladder chalone C has enzyme linked immunosorbent assay, colloidal gold method, Enzyme-linked Immunosorbent Assay at present
Method operation is complicated, the longest, and has certain professional skill requirement to operator, and cannot detection by quantitative, colloidal gold method can only
Qualitative detection.
Summary of the invention
The technical problem to be solved be in order to overcome operation complicated and can not the defect of detection by quantitative, and provide
A kind of test kit measuring bladder chalone C and preparation method thereof.
The technical scheme that the present invention solves above-mentioned technical problem and provides is: the invention discloses and a kind of measures bladder chalone C
Test kit, forms test kit including reagent R1 independent of each other and reagent R2 biliquid component, including composition and corresponding content
For:
Reagent R1:
Buffer 40 ~ 180 mmol/L
Inorganic ion 50 ~ 100 mmol/L
Accelerator 5 ~ 20 g/L
Stabilizer 5 ~ 15 g/L
Preservative 0.4 ~ 1.2 g/L
Its solvent is purified water
Reagent R2:
Buffer 40 ~ 180 mmol/L
Stabilizer 10 ~ 30 g/L
Suspending agent 5 ~ 35 mmol/L
Surfactant 0.5 ~ 2.0 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.2 ~ 1.2 g/L
Preservative 0.4 ~ 1.2 g/L
Latex is coated anti-human bladder chalone C antibody 0.5 ~ 3.0 g/L
Its solvent is purified water.
As preferably, the invention discloses a kind of test kit measuring bladder chalone C, including reagent R1 independent of each other and examination
Agent R2 biliquid component composition test kit, including composition and corresponding content be:
Reagent R1:
Buffer 110 mmol/L
Inorganic ion 70 mmol/L
Accelerator 12 g/L
Stabilizer 10 g/L
Preservative 0.8 g/L
Its solvent is purified water
Reagent R2:
Buffer 110 mmol/L
Stabilizer 20 g/L
Suspending agent 20 mmol/L
Surfactant 1.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.7 g/L
Preservative 0.8 g/L
Latex is coated anti-human bladder chalone C antibody 2.0 g/L
Its solvent is purified water.
As preferably, in described reagent R1, described buffer uses MES buffer, MOPS buffer, MOPSO to delay
Rush the combination of one or more in liquid, HEPES buffer, Tris buffer, phosphate buffer, described inorganic ion
The combination of one or more in employing sodium chloride, potassium chloride, magnesium chloride, magnesium sulfate, described accelerator employing Polyethylene Glycol-
2000, the combination of one or more in PEG-4000, PEG-8 000, polyvinylpyrrolidone, described is steady
Determine agent and use the combination of one or more in bovine serum albumin, mannose, sorbitol, disodiumedetate, fructose,
Described preservative uses the combination of one or more in phenol, sodium benzoate, sodium azide.
As preferably, in described reagent R2, described buffer uses MES buffer, MOPS buffer, MOPSO to delay
Rushing the combination of one or more in liquid, HEPES buffer, Tris buffer, phosphate buffer, described stabilizer uses
The combination of one or more in bovine serum albumin, mannose, sorbitol, disodiumedetate, fructose, described helps
Suspension uses one or more in arabic gum, sodium alginate, silica sol, glycerol to combine, described surface
Activating agent uses triton x-100, described preservative to use the group of one or more in phenol, sodium benzoate, sodium azide
Close.
As preferably, in described reagent R2, described latex is coated the preparation method of anti-human bladder chalone C antibody and is: with 50
The polystyrene microsphere dilution that particle diameter is 30 ~ 200nm is become the matter of contained polystyrene microsphere by the MES buffer of mmol/L
Amount concentration is the solution of 1.5%, then adds the 1-ethyl-3-(3-DimethylAminopropyl of 0.8 mg in every milliliter of solution) carbonization
Diimmonium salt hydrochlorate, reacts 2 hours under conditions of 20 DEG C ~ 37 DEG C, uses centrifuge, under the rotating speed of 20000 rpm/min
Centrifugal 30 minutes, remove supernatant, then precipitation is suspended in the MES buffer of 50 mmol/L, ultrasonic disperse, re-uses centrifugal
Machine, is centrifuged 30 minutes under the rotating speed of 20000 rpm/min, removes supernatant, then precipitation is suspended in the MES buffering of 50 mmol/L
In liquid so that the final mass concentration of the polystyrene microsphere in solution is 3.0%, ultrasonic disperse, then limit dispersion limit addition etc.
The MES buffer of the anti-human bladder chalone C antibody containing 8/ml of volume, mix and blend, reacts 2 under conditions of 20 DEG C ~ 37 DEG C
Hour, till the final mass concentration of the polystyrene microsphere in solution is 1.5%, add bovine serum albumin so that cattle
Till sero-abluminous ultimate density is 10 g/L, closes 12 ~ 18 hours under the conditions of 4 DEG C, latex bag can be prepared
By anti-human bladder chalone C antibody.
As preferably, described anti-human bladder chalone C antibody be containing can be specific binding with human cystatin C Fab function
The complete antibody at position or antibody fragment.
As preferably, the invention also discloses preparation method and the using method of the test kit of said determination bladder chalone C, bag
Include following steps:
A () prepares reagent according to following component content:
The invention discloses a kind of test kit measuring bladder chalone C, including reagent R1 independent of each other and reagent R2 biliquid group
Be grouped into test kit, including composition and corresponding content be:
Reagent R1:
Buffer 40 ~ 180 mmol/L
Inorganic ion 50 ~ 100 mmol/L
Accelerator 5 ~ 20 g/L
Stabilizer 5 ~ 15 g/L
Preservative 0.4 ~ 1.2 g/L
Its solvent is purified water
Reagent R2:
Buffer 40 ~ 180 mmol/L
Stabilizer 10 ~ 30 g/L
Suspending agent 5 ~ 35 mmol/L
Surfactant 0.5 ~ 2.0 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.2 ~ 1.2 g/L
Preservative 0.4 ~ 1.2 g/L
Latex is coated anti-human bladder chalone C antibody 0.5 ~ 3.0 g/L
Its solvent is purified water；
B sample to be tested is mixed by () with reagent R1 and reagent R2 so that it is fully react；
C () measures reacted absorbance difference with automatic clinical chemistry analyzer；
D () calculates the concentration of bladder chalone C in sample according to absorbance changing value.
As preferably, in step (b), the volume ratio of described reagent R1 and reagent R2 is 9:2.
As preferably, in step (b), the volume ratio of the cumulative volume of described sample to be tested and reagent R1 and reagent R2 is 1:
Between 5 to 1:100.
The Cleaning Principle of the present invention is: in sample, bladder chalone C can specificity corresponding with reagent anti-Cys-C antibody knot
Closing and form antigen-antibody complex, produce certain turbidity, this turbidity height content with antigen in the presence of certain antibody just becomes
Ratio, is measured turbidity under certain wavelength and can be carried out the quantitative determination of bladder chalone C by multiple spot calibration curve.
Bladder chalone C (Cys-C) activity (mg/L)=C in sampleS ×(mg/L)
In formula: Δ ATThe sample cell absorbance compared with blank tube absorbance
ΔASThe calibration pipe absorbance compared with blank tube absorbance
CSThe concentration of Cys-C in calibration solution
Compared with prior art, the present invention has following advantageous benefits: the present invention with the addition of glycerol as suspending in reagent R2
Agent, under the effect of surfactant so that latex be coated anti-human bladder chalone C antibody can even suspension in reagent R2, simultaneously
With the addition of polyoxyethylene polyoxypropylene copolymer in reagent R2, this copolymer can make the most mutually to assemble between latex particle, greatly
Improve greatly the stability of reagent R2 and the titer of antibody, when participating in reaction after adding sample, accelerator in reagent R1
Under effect, anti-human bladder chalone C antibody can be coated rapidly with latex and react, suspend because latex is coated anti-human bladder chalone C antibody
It is uniformly dispersed, even if under relatively low concentration, it is possible to react rapidly so that the detection sensitivity of reagent carries the most further
Height, the accuracy of detection is also considerably improved, and easy to operate.
Detailed description of the invention
Further illustrate below in conjunction with specific embodiment, but the present invention is not limited in these embodiments.
Embodiment 1
The test kit of the present invention includes reagent R1 independent of each other and reagent R2 biliquid component, wherein
Reagent R1:
Tris buffer 110 mmol/L
Potassium chloride 70 mmol/L
PEG-4000 12 g/L
Bovine serum albumin 10 g/L
Phenol 0.8 g/L
Its solvent is purified water
Reagent R2:
MES buffer 110 mmol/L
Mannose 20 g/L
Sodium alginate 20 mmol/L
Triton x-100 1.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.7 g/L
Phenol 0.8 g/L
Latex is coated anti-human bladder chalone C antibody 2.0 g/L
Its solvent is purified water.
Embodiment 2
The test kit of the present invention includes reagent R1 independent of each other and reagent R2 biliquid component, wherein
Reagent R1:
MOPS buffer 40 mmol/L
Sodium chloride 100 mmol/L
PEG-8 000 20 g/L
Sorbitol 5 g/L
Sodium benzoate 0.4 g/L
Its solvent is purified water
Reagent R2:
MOPS buffer 180 mmol/L
Bovine serum albumin 10 g/L
Sodium alginate 35 mmol/L
Triton x-100 0.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 1 .2 g/L
Sodium azide 0.4 g/L
Latex is coated anti-human bladder chalone C antibody 3.0 g/L
Its solvent is purified water.
Embodiment 3
The preparation and application of test kit
1, latex is coated the preparation of anti-human bladder chalone C antibody: be the polyphenyl of 120nm with the MES buffer of 50 mmol/L by particle diameter
The dilution of ethylene microsphere becomes the solution that mass concentration is 1.5% of contained polystyrene microsphere, then adds in every milliliter of solution
1-ethyl-3-(3-the DimethylAminopropyl of 0.8 mg) carbodiimide hydrochloride, reacts 2 hours under conditions of 25 DEG C, makes
With centrifuge, it is centrifuged 30 minutes under the rotating speed of 20000 rpm/min, removes supernatant, then precipitation is suspended in 50 mmol/L's
In MES buffer, ultrasonic disperse, re-use centrifuge, be centrifuged 30 minutes under the rotating speed of 20000 rpm/min, remove supernatant, then
Precipitation is suspended in the MES buffer of 50 mmol/L so that the final mass concentration of the polystyrene microsphere in solution is
3.0%, ultrasonic disperse, then dispersion limit in limit adds the MES buffer of isopyknic anti-human bladder chalone C antibody containing 8/ml, mixed
Close stirring, react 2 hours under conditions of 25 DEG C, until the final mass concentration of the polystyrene microsphere in solution is 1.5% to be
Only, add bovine serum albumin so that till the ultimate density of bovine serum albumin is 10 g/L, under the conditions of 4 DEG C, close 15
Hour, latex can be prepared and be coated anti-human bladder chalone C antibody；
2, reagent is prepared according to following component content:
Reagent R1:
Tris buffer 110 mmol/L
Potassium chloride 70 mmol/L
PEG-4000 12 g/L
Bovine serum albumin 10 g/L
Phenol 0.8 g/L
Its solvent is purified water
Reagent R2:
MES buffer 110 mmol/L
Mannose 20 g/L
Sodium alginate 20 mmol/L
Triton x-100 1.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.7 g/L
Phenol 0.8 g/L
Latex is coated anti-human bladder chalone C antibody 2.0 g/L
Its solvent is purified water；
3, automatic clinical chemistry analyzer parameter is arranged
(a) detection temperature: 37 DEG C；
(b) detection wavelength: dominant wavelength 546nm, commplementary wave length 800nm；
(c) response time: 10min, wherein, incubation time 5min, measure immediately after adding reagent R2 and read absorbance
Read absorbance A 2 after A1,5min, calculate absorbance changes delta A=A2-A1；
(d) the Direction of Reaction: positive reaction；
4, detecting step
A () takes 225 μ l reagent R1 and the mixing of 3 μ l samples to be tested；
B solution after mixing is hatched 5min under conditions of 37 DEG C by ()；
C () adds 50 μ l reagent R2, measure immediately and read absorbance A 1, reads absorbance A 2 after 5min, calculates absorbance change
Δ A=A2-A1；
D () is according to bladder chalone C in sample (Cys-C) activity (mg/L)=CS × (mg/L) Guang in sample is calculated
The concentration of chalone C.
Embodiment 4
The preparation and application of test kit
1, latex is coated the preparation of anti-human bladder chalone C antibody: be the polyphenyl of 70nm with the MES buffer of 50 mmol/L by particle diameter
The dilution of ethylene microsphere becomes the solution that mass concentration is 1.5% of contained polystyrene microsphere, then adds in every milliliter of solution
1-ethyl-3-(3-the DimethylAminopropyl of 0.8 mg) carbodiimide hydrochloride, reacts 2 hours under conditions of 25 DEG C, makes
With centrifuge, it is centrifuged 30 minutes under the rotating speed of 20000 rpm/min, removes supernatant, then precipitation is suspended in 50 mmol/L's
In MES buffer, ultrasonic disperse, re-use centrifuge, be centrifuged 30 minutes under the rotating speed of 20000 rpm/min, remove supernatant, then
Precipitation is suspended in the MES buffer of 50 mmol/L so that the final mass concentration of the polystyrene microsphere in solution is
3.0%, ultrasonic disperse, then dispersion limit in limit adds the MES buffer of isopyknic anti-human bladder chalone C antibody containing 8/ml, mixed
Close stirring, react 2 hours under conditions of 25 DEG C, until the final mass concentration of the polystyrene microsphere in solution is 1.5% to be
Only, add bovine serum albumin so that till the ultimate density of bovine serum albumin is 10 g/L, under the conditions of 4 DEG C, close 15
Hour, latex can be prepared and be coated anti-human bladder chalone C antibody；
2, reagent is prepared according to following component content:
Reagent R1:
MOPS buffer 40 mmol/L
Sodium chloride 100 mmol/L
PEG-8 000 20 g/L
Sorbitol 5 g/L
Sodium benzoate 0.4 g/L
Its solvent is purified water
Reagent R2:
MOPS buffer 180 mmol/L
Bovine serum albumin 10 g/L
Sodium alginate 35 mmol/L
Triton x-100 0.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 1 .2 g/L
Sodium azide 0.4 g/L
Latex is coated anti-human bladder chalone C antibody 3.0 g/L
Its solvent is purified water；
3, automatic clinical chemistry analyzer parameter is arranged
(a) detection temperature: 37 DEG C；
(b) detection wavelength: dominant wavelength 546nm, commplementary wave length 800nm；
(c) response time: 10min, wherein, incubation time 5min, measure immediately after adding reagent R2 and read absorbance
Read absorbance A 2 after A1,5min, calculate absorbance changes delta A=A2-A1；
(d) the Direction of Reaction: positive reaction；
4, detecting step
A () takes 225 μ l reagent R1 and the mixing of 3 μ l samples to be tested；
B solution after mixing is hatched 5min under conditions of 37 DEG C by ()；
C () adds 50 μ l reagent R2, measure immediately and read absorbance A 1, reads absorbance A 2 after 5min, calculates absorbance change
Δ A=A2-A1；
D () is according to bladder chalone C in sample (Cys-C) activity (mg/L)=CS × (mg/L) Guang in sample is calculated
The concentration of chalone C.
The table 1 test kit measuring bladder chalone C obtained by embodiment 1 and the mensuration bladder chalone C obtained by embodiment 2
The result that respectively quality-control product 1 is measured of test kit, wherein the concentration of the bladder chalone C in quality-control product 1 is 1.00 mg/L,
Measurement result is shown in Table 1:
Table 1
| 1st time (mg/L) | 2nd time (mg/L) | 3rd time (mg/L) | Average (mg/L) | Deviation (%) | |
| Embodiment 1 | 0.98 | 1.02 | 0.98 | 0.99 | 1.00 |
| Embodiment 2 | 1.02 | 1.03 | 1.03 | 1.03 | 3.00 |
As shown in Table 1, the test kit measuring bladder chalone C obtained by the present invention is less to the measurement result deviation of quality-control product 1, because of
This accuracy of measurement is high, and embodiment 1 is optimum selection.
The table 2 test kit measuring bladder chalone C obtained by embodiment 1 and the examination measuring bladder chalone C obtained by embodiment 2
The result that quality-control product 2 is measured by agent box respectively, wherein the concentration of the bladder chalone C in quality-control product 2 is 2.40 mg/L, measures
The results are shown in Table 2:
Table 2
| 1st time (mg/L) | 2nd time (mg/L) | 3rd time (mg/L) | Average (mg/L) | Deviation (%) | |
| Embodiment 1 | 2.37 | 2.39 | 2.48 | 2.41 | 0.42 |
| Embodiment 2 | 2.32 | 2.36 | 2.35 | 2.34 | 2.50 |
As shown in Table 2, the test kit measuring bladder chalone C obtained by the present invention is less to the measurement result deviation of quality-control product 2, because of
This accuracy of measurement is high, and embodiment 1 is optimum selection.
The table 3 test kit measuring bladder chalone C obtained by embodiment 3 is surveyed the most repeatedly to what same sample to be tested was carried out
The test kit measuring bladder chalone C calmly and obtained by embodiment 4 is repeatedly repeatedly measured what same sample to be tested was carried out, to institute
The result obtained carries out the calculating of SD and CV, and result is as follows:
Table 3
The precision of the test kit measuring bladder chalone C obtained by the present invention is relatively good as shown in Table 3, and as shown in Table 1, real
Executing example 3 is optimum selection.
The principle of above-described embodiment only illustrative present invention and effect thereof, not for limiting the present invention.Any ripe
Above-described embodiment all can be modified under the spirit and the scope of the present invention or change by the personage knowing this technology.Cause
This, have usually intellectual such as complete with institute under technological thought without departing from disclosed spirit in art
All equivalences become are modified or change, and must be contained by the claim of the present invention.
Claims (9)
1. the test kit measuring bladder chalone C, it is characterised in that: include reagent R1 independent of each other and reagent R2 biliquid group
Be grouped into test kit, including composition and corresponding content be:
Reagent R1:
Buffer 40 ~ 180 mmol/L
Inorganic ion 50 ~ 100 mmol/L
Accelerator 5 ~ 20 g/L
Stabilizer 5 ~ 15 g/L
Preservative 0.4 ~ 1.2 g/L
Its solvent is purified water
Reagent R2:
Buffer 40 ~ 180 mmol/L
Stabilizer 10 ~ 30 g/L
Suspending agent 5 ~ 35 mmol/L
Surfactant 0.5 ~ 2.0 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.2 ~ 1.2 g/L
Preservative 0.4 ~ 1.2 g/L
Latex is coated anti-human bladder chalone C antibody 0.5 ~ 3.0 g/L
Its solvent is purified water.
A kind of test kit measuring bladder chalone C the most according to claim 1, it is characterised in that: include examination independent of each other
Agent R1 and reagent R2 biliquid component composition test kit, including composition and corresponding content be:
Reagent R1:
Buffer 110 mmol/L
Inorganic ion 70 mmol/L
Accelerator 12 g/L
Stabilizer 10 g/L
Preservative 0.8 g/L
Its solvent is purified water
Reagent R2:
Buffer 110 mmol/L
Stabilizer 20 g/L
Suspending agent 20 mmol/L
Surfactant 1.5 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.7 g/L
Preservative 0.8 g/L
Latex is coated anti-human bladder chalone C antibody 2.0 g/L
Its solvent is purified water.
A kind of test kit measuring bladder chalone C the most according to claim 1 and 2, it is characterised in that: described reagent R1
In, described buffer uses MES buffer, MOPS buffer, MOPSO buffer, HEPES buffer, Tris buffer, phosphorus
The combination of one or more in phthalate buffer, described inorganic ion uses sodium chloride, potassium chloride, magnesium chloride, sulphuric acid
The combination of one or more in magnesium, described accelerator use Polyethylene glycol-2000, PEG-4000, Polyethylene Glycol-
8000, the combination of one or more in polyvinylpyrrolidone, described stabilizer use bovine serum albumin, mannose,
The combination of one or more in sorbitol, disodiumedetate, fructose, described preservative uses phenol, benzoic acid
The combination of one or more in sodium, sodium azide.
A kind of test kit measuring bladder chalone C the most according to claim 1 and 2, it is characterised in that: described reagent R2
In, described buffer uses MES buffer, MOPS buffer, MOPSO buffer, HEPES buffer, Tris buffer, phosphorus
The combination of one or more in phthalate buffer, described stabilizer uses bovine serum albumin, mannose, sorbitol, second
The combination of one or more in edetate disodium, fructose, described suspending agent uses arabic gum, sodium alginate, glue
One or more in body silicon dioxide, glycerol combine, and described surfactant uses triton x-100, described
Preservative uses the combination of one or more in phenol, sodium benzoate, sodium azide.
A kind of test kit measuring bladder chalone C the most according to claim 1 and 2, it is characterised in that: described reagent R2
In, described latex is coated the preparation method of anti-human bladder chalone C antibody and is: be 30 with the MES buffer of 50 mmol/L by particle diameter
The polystyrene microsphere dilution of ~ 200nm becomes the solution that mass concentration is 1.5% of contained polystyrene microsphere, then every milli
Rise the 1-ethyl-3-(3-DimethylAminopropyl adding 0.8 mg in solution) carbodiimide hydrochloride, at 20 DEG C ~ 37 DEG C
Under the conditions of react 2 hours, use centrifuge, under the rotating speed of 20000 rpm/min centrifugal 30 minutes, remove supernatant, then will be heavy
Form sediment in the MES buffer being suspended in 50 mmol/L, ultrasonic disperse, re-use centrifuge, under the rotating speed of 20000 rpm/min
Centrifugal 30 minutes, remove supernatant, then precipitation is suspended in the MES buffer of 50 mmol/L so that the polystyrene in solution is micro-
The final mass concentration of ball is 3.0%, ultrasonic disperse, and then dispersion limit in limit adds isopyknic anti-human bladder chalone C containing 8/ml
The MES buffer of antibody, mix and blend, reacts 2 hours, until the polystyrene in solution is micro-under conditions of 20 DEG C ~ 37 DEG C
Till the final mass concentration of ball is 1.5%, add bovine serum albumin so that the ultimate density of bovine serum albumin is 10 g/
Till L, close 12 ~ 18 hours under the conditions of 4 DEG C, latex can be prepared and be coated anti-human bladder chalone C antibody.
A kind of test kit measuring bladder chalone C the most according to claim 5, it is characterised in that: described anti-human bladder chalone C
Antibody is the complete antibody containing Fab functional part that can be specific binding with human cystatin C or antibody fragment.
The preparation method of a kind of test kit measuring bladder chalone C the most according to claim 1 and 2 and using method, it is special
Levy and be: comprise the following steps:
A () prepares reagent according to following component content:
The invention discloses a kind of test kit measuring bladder chalone C, including reagent R1 independent of each other and reagent R2 biliquid group
Be grouped into test kit, including composition and corresponding content be:
Reagent R1:
Buffer 40 ~ 180 mmol/L
Inorganic ion 50 ~ 100 mmol/L
Accelerator 5 ~ 20 g/L
Stabilizer 5 ~ 15 g/L
Preservative 0.4 ~ 1.2 g/L
Its solvent is purified water
Reagent R2:
Buffer 40 ~ 180 mmol/L
Stabilizer 10 ~ 30 g/L
Suspending agent 5 ~ 35 mmol/L
Surfactant 0.5 ~ 2.0 mL/L
Polyoxyethylene polyoxypropylene copolymer 0.2 ~ 1.2 g/L
Preservative 0.4 ~ 1.2 g/L
Latex is coated anti-human bladder chalone C antibody 0.5 ~ 3.0 g/L
Its solvent is purified water；
B sample to be tested is mixed by () with reagent R1 and reagent R2 so that it is fully react；
C () measures reacted absorbance difference with automatic clinical chemistry analyzer；
D () calculates the concentration of bladder chalone C in sample according to absorbance changing value.
The preparation method of a kind of test kit measuring bladder chalone C the most according to claim 7 and using method, its feature exists
In: in step (b), the volume ratio of described reagent R1 and reagent R2 is 9:2.
The preparation method of a kind of test kit measuring bladder chalone C the most according to claim 7 and using method, its feature exists
In: in step (b), the volume ratio of the cumulative volume of described sample to be tested and reagent R1 and reagent R2 is between 1:5 to 1:100.
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| CN201610376097.5A CN106093426A (en) | 2016-05-31 | 2016-05-31 | A kind of test kit measuring bladder chalone C and preparation method thereof |
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| CN106896227A (en) * | 2017-04-05 | 2017-06-27 | 济南蓄琪生物技术有限公司 | A kind of myeloperoxidase enzyme detection kit |
| CN107228940A (en) * | 2017-05-31 | 2017-10-03 | 吉林省汇酉生物技术股份有限公司 | The detection reagent and method of a kind of bladder chalone C |
| CN107607707A (en) * | 2017-03-31 | 2018-01-19 | 迈克生物股份有限公司 | Suppress the bladder chalone C latex enhancing immune of rheumatoid factor interference than turbid kit |
| CN107677841A (en) * | 2017-09-30 | 2018-02-09 | 安徽伊普诺康生物技术股份有限公司 | A kind of preparation method of NBAP detection kit |
| CN107677804A (en) * | 2017-09-30 | 2018-02-09 | 安徽伊普诺康生物技术股份有限公司 | A kind of NBAP detection kit and its application method |
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| CN108872604A (en) * | 2018-07-27 | 2018-11-23 | 金华市强盛生物科技有限公司 | A kind of cystatin C detection kit |
| CN109358009A (en) * | 2018-10-26 | 2019-02-19 | 武汉百德瑞康生物技术有限公司 | Bladder chalone C determining reagent kit and preparation method thereof and detection method |
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| CN109813908A (en) * | 2018-12-29 | 2019-05-28 | 宁波普瑞柏生物技术股份有限公司 | Eliminate the method and kit of hook effect in cystatin C detection |
| CN112698023A (en) * | 2020-12-01 | 2021-04-23 | 三诺生物传感股份有限公司 | Preserving fluid of alkaline phosphatase labeled conjugate and preparation method thereof |
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| CN107607707A (en) * | 2017-03-31 | 2018-01-19 | 迈克生物股份有限公司 | Suppress the bladder chalone C latex enhancing immune of rheumatoid factor interference than turbid kit |
| CN107607707B (en) * | 2017-03-31 | 2020-04-07 | 迈克生物股份有限公司 | Cystatin C latex enhanced immunoturbidimetry kit for inhibiting rheumatoid factor interference |
| CN106896227A (en) * | 2017-04-05 | 2017-06-27 | 济南蓄琪生物技术有限公司 | A kind of myeloperoxidase enzyme detection kit |
| CN107228940B (en) * | 2017-05-31 | 2019-09-24 | 吉林省汇酉生物技术股份有限公司 | A kind of detection reagent and method of cystatin C |
| CN107228940A (en) * | 2017-05-31 | 2017-10-03 | 吉林省汇酉生物技术股份有限公司 | The detection reagent and method of a kind of bladder chalone C |
| CN107677841A (en) * | 2017-09-30 | 2018-02-09 | 安徽伊普诺康生物技术股份有限公司 | A kind of preparation method of NBAP detection kit |
| CN107677804A (en) * | 2017-09-30 | 2018-02-09 | 安徽伊普诺康生物技术股份有限公司 | A kind of NBAP detection kit and its application method |
| CN107727846A (en) * | 2017-09-30 | 2018-02-23 | 安徽伊普诺康生物技术股份有限公司 | A kind of preparation method of the Complement C4 detection kit of stabilization |
| CN107741499A (en) * | 2017-09-30 | 2018-02-27 | 安徽伊普诺康生物技术股份有限公司 | The Complement C4 detection kit and its application method of a kind of stabilization |
| CN107741491A (en) * | 2017-09-30 | 2018-02-27 | 安徽伊普诺康生物技术股份有限公司 | A kind of kit for determining urine galactose and its preparation application method |
| CN108107201A (en) * | 2017-12-20 | 2018-06-01 | 青岛汉唐生物科技有限公司 | A kind of cystatin C detection kit and preparation method thereof |
| CN108872604A (en) * | 2018-07-27 | 2018-11-23 | 金华市强盛生物科技有限公司 | A kind of cystatin C detection kit |
| CN109358009A (en) * | 2018-10-26 | 2019-02-19 | 武汉百德瑞康生物技术有限公司 | Bladder chalone C determining reagent kit and preparation method thereof and detection method |
| CN109358009B (en) * | 2018-10-26 | 2021-08-10 | 武汉百德瑞康生物技术有限公司 | Cystatin C determination kit, preparation method and detection method thereof |
| CN109613237A (en) * | 2018-12-28 | 2019-04-12 | 上海高踪医疗器械科技有限公司 | A kind of cystatin C detection kit |
| CN109813908A (en) * | 2018-12-29 | 2019-05-28 | 宁波普瑞柏生物技术股份有限公司 | Eliminate the method and kit of hook effect in cystatin C detection |
| CN112698023A (en) * | 2020-12-01 | 2021-04-23 | 三诺生物传感股份有限公司 | Preserving fluid of alkaline phosphatase labeled conjugate and preparation method thereof |
| CN112698023B (en) * | 2020-12-01 | 2023-12-15 | 三诺生物传感股份有限公司 | Preservation solution of alkaline phosphatase-labeled conjugate and preparation method thereof |
| CN115541895A (en) * | 2022-11-29 | 2022-12-30 | 天津德祥生物技术股份有限公司 | Formula liquid for improving sensitivity of micro-fluidic inverse detection card and application |
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