CN108680751A - Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein - Google Patents
Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein Download PDFInfo
- Publication number
- CN108680751A CN108680751A CN201810298348.1A CN201810298348A CN108680751A CN 108680751 A CN108680751 A CN 108680751A CN 201810298348 A CN201810298348 A CN 201810298348A CN 108680751 A CN108680751 A CN 108680751A
- Authority
- CN
- China
- Prior art keywords
- antibody
- saa
- protein
- preparation
- pbs
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
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
- G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
-
- 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
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
-
- 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
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/46—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans from vertebrates
- G01N2333/47—Assays involving proteins of known structure or function as defined in the subgroups
Abstract
Present invention is disclosed a kind of preparation methods of the time-resolved fluoroimmunoassay chromatograph test strip of quantitatively detection SAA, including：Nitrocellulose filter is coated with detection line and nature controlling line, and detection line detects antibody using SAA, and nature controlling line captures the secondary antibody of antibody using anti-SAA；Prepare bonding pad comprising：In PBS containing the enhanced time-resolved fluorescence microballoon of nanometer, EDC and NHS, centrifugation is added, precipitation is dissolved in after PBS is washed in PBS；SAA captures antibody and uses cleaning solution centrifuge washing；SAA capture antibody after washing is mixed with the PB dissolved with fluorescent microsphere, so that SAA capture antibody is coupled with fluorescent microsphere, the mixed liquor after coupling is sprayed at carrier.
Description
Technical field
The present invention relates to field of clinical immunology, and in particular to it is a kind of quantitatively detection serum amyloid A protein when
Between resolved fluorometric immuno-chromatographic test paper strip and preparation method thereof.
Background technology
Serum amyloid A protein (serum amyloid A, SAA) is a kind of acute time limit reactive protein, belongs to and carries fat egg
Heterogeneous proteinoid in white family, relative molecular weight about 12000D.In the acute time limit reacts, pierced through IL-1, IL-6 and TNF
Swash, SAA is synthesized in liver by the macrophage and fibroblast that are activated, can be increased to the 100-1000 of initial concentration
Times, but half-life period is extremely short, and only 50 minutes or so.SAA is related with high-density lipoprotein (HDL), it can be adjusted during inflammation
The metabolism of high-density lipoprotein.Mono- especially important characteristic of SAA is that its catabolite can be with amyloid A (AA) fibrinogen
Mode be deposited in different organs, this is a kind of serious complication in chronic inflammatory diseases.
Similar with c reactive protein (CRP), the content concn of SAA is the sensitive indicator for reflecting infectious diseases Earlier period of inflammation,
Help to diagnose inflammation, assesses its activity, monitors its activity and treatment.But in diagnosis virus infection, kidney occur for SAA detections
The patient (patient for especially carrying out immunosuppressive therapy) of graft-rejection and the capsule with adrenocortical hormones in treating
Property fibrosis patients in terms of, it is more conclusive than c reactive protein.The study found that in the case for suffering from inflammatory arthritis, SAA and disease
The relationship of activity is most close.It detects c reactive protein simultaneously and SAA can improve diagnostic sensitivity to infection.For amyloid
SAA levels are returned back to the normally treatment for objective, can improve the state of an illness by albumin A amyloidosis patient.
Detection for SAA, method used at present are mainly chemoluminescence method, immunoturbidimetry, colloidal gold etc., but this
A little methods have the characteristics that respective and deficiency.Chemiluminescence method needs large-scale instrument, and takes for a long time, with high costs；It is immune
It is lower than turbid sensitivity, poor accuracy；Colloidal gold method cannot be quantified and be detected.
Invention content
The purpose of the present invention is to provide a kind of highly sensitive, high specific SAA clinic quantitative testing test paper items.
For achieving the above object, the present invention provides a kind of time-resolved fluoroimmunoassay chromatography examination of quantitatively detection SAA
The preparation method of paper slip, including：
Nitrocellulose filter is coated with detection line and nature controlling line, and the detection line detects antibody using SAA, and the nature controlling line makes
The secondary antibody of antibody is captured with anti-SAA；
Prepare bonding pad comprising：
In PBS containing the enhanced time-resolved fluorescence microballoon of nanometer, the EDC and NHS of final concentration of 10mg/ml is added,
Centrifugation, precipitation are dissolved in after PBS is washed in PBS；
SAA captures antibody and uses antibody cleaning solution centrifuge washing；
By after washing SAA capture antibody mixed with the PBS dissolved with fluorescent microsphere so that SAA capture antibody and
Fluorescent microsphere is coupled；
Mixed liquor after coupling is sprayed into carrier；
Wherein, PBS PH6-7, concentration 0.05mol/L.
As being further improved for an embodiment of the present invention, a diameter of 100nm-300nm of fluorescent microsphere.
As being further improved for an embodiment of the present invention, the weight ratio that SAA capture antibody is mixed with fluorescent microsphere is
1/10-1/20。
As being further improved for an embodiment of the present invention, after SAA captures antibody and fluorescent microsphere coupling, through confining liquid
Closing, then sprays to carrier, and the confining liquid contains the 35 and a concentration of 10- of card pool that mass percent is 0.1-1%
The glycine of 50mmol/ml.
As being further improved for an embodiment of the present invention, the 0.05mol/L PB that antibody cleaning solution is PH7-8 are buffered
Liquid.
As being further improved for an embodiment of the present invention, carrier is glass fibre membrane.
Further include step after antibody is mixed with fluorescent microsphere as being further improved for an embodiment of the present invention：
The pH value that the SAA is captured to the mixed liquor of antibody and fluorescent microsphere is adjusted to 7.2 ± 0.2.
As being further improved for an embodiment of the present invention, the SAA detection antibody dosages of each detection line are 0.5-
2mg。
As being further improved for an embodiment of the present invention, the secondary antibody of the anti-SAA capture antibody of each nature controlling line, dosage
For 0.5-2mg.
To achieve the above object, an embodiment of the present invention provides a kind of time-resolved fluoroimmunoassay quantitatively detecting SAA
Chromatograph test strip is made of bottom plate, sample pad, bonding pad, nitrocellulose filter and blotting paper, and test strips are by above-mentioned method system
.
Compared with prior art, the present invention is by introducing double antibody sandwich method and time-resolved fluoroimmunoassay chromatographic technique
In the detection of SAA, in conjunction with fluorescence detector, realize the quantitative detection of SAA, and high sensitivity, batch in, difference between batch it is small, to face
Bed use provides great convenience.
Description of the drawings
Fig. 1 is the canonical plotting of the embodiment of the present invention 1；
Fig. 2 is the canonical plotting of the embodiment of the present invention 2；
Fig. 3 is the test strips that SAA is quantitatively detected made from embodiment 1 and the contrast test results relevance of immunoturbidimetry
Analysis；
Fig. 4 is the test strips that SAA is quantitatively detected made from embodiment 2 and the contrast test results relevance of immunoturbidimetry
Analysis.
Specific implementation mode
Below with reference to specific implementation mode shown in the drawings, the present invention will be described in detail.But these embodiments are simultaneously
The present invention is not limited, structure that those skilled in the art are made according to these embodiments, method or functionally
Transformation is included within the scope of protection of the present invention.
Test strips provided by the invention get stuck by plastics, bottom plate, sample pad, bonding pad, nitrocellulose filter (NC films) and
Blotting paper forms.Sample pad is blank glass fiber film.The enhanced time-resolved fluorescence microballoon mark of nanometer is coated on bonding pad
The SAA capture antibody (capture antibody) of note.Detection band and quality control band are coated on NC films, wherein detection band is fixed
There is the SAA detection antibody (test antibody) for identifying different epitopes, quality control band is fixed with can be anti-in conjunction with anti-SAA captures
The secondary antibody of body.
Bonding pad is glass fibre membrane, coats the enhanced time-resolved fluorescence microballoon of nanometer for being connected with antibody thereon.
Coated detection band and quality control band are mutually parallel on nitrocellulose filter (NC films), and keep between each other certain
Interval.
The enhanced time-resolved fluorescence microballoon of nanometer selects the microballoon of diameter 100nm-300nm.
In test strips get stuck loaded on plastics, assembled in environment of the humidity less than 35%.
The test strips of the present invention are made by following steps：
Detection line and nature controlling line are prepared on NC films, detection line is that SAA detects antibody (test antibody), and nature controlling line is
The secondary antibody of anti-SAA capture antibody can be combined；
Spraying is connected with the nanometer enhanced time point of SAA capture antibody (capture antibody) on glass fibre membrane
It distinguishes fluorescent microsphere, bonding pad is made；
The strip that the glass fibre membrane of one blank is cut into suitable size, as sample pad；
Bottom plate, NC films, blotting paper, bonding pad, sample pad are staggeredly pasted successively, cut into corresponding size, are packed into plastic clip
Shell.
Wherein, it is coated with detection line on NC films, the method for nature controlling line includes the following steps：
Detection line detects antibody using SAA, and dosage is per detection line 0.5-2mg.Antibody uses diluted to concentration
It is used after range, dilution selects PBS+ (1-10) % trehaloses+(0-5) %NaCl.
For nature controlling line using the secondary antibody of anti-SAA capture antibody, dosage is per nature controlling line 0.5-2mg.Higher than the antibody of this concentration
Using being used after diluted to concentration range, dilution selects PBS+ (1-10) % trehaloses+(0-5) %NaCl.
Using quantitative spray film instrument with the concentration of 0.8-1.5ul/cm by above-mentioned two kinds of antibody graduation on NC films, keep two lines
It is spaced 0.5cm or more.
25-60 DEG C dries 1-5 hours, kept dry.
The production method of bonding pad includes the following steps：
EDC, NHS of final concentration of 10mg/ml, room are added in PBS containing the enhanced time-resolved fluorescence microballoon of nanometer
Temperature reaction 30min-2 hours, wherein PBS PH6-7, concentration 0.05mol/L；
15000-25000rpm/min centrifuges 10min, and supernatant, PBS is gone to wash 2 times, be dissolved in PBS；
SAA is captured antibody and is washed 3 times using ultra-filtration centrifuge tube, and cleaning solution is the 0.05mol/L PB buffer solutions of PH7-8；
Capture antibody after washing is mixed into coupling with fluorescent microsphere, control antibody and fluorescent microsphere weight ratio are 1/10-
1/20, supplementing suitable PB buffer solutions makes PH be equal to 7.2 ± 0.2, mixing；
Room temperature reaction 1-4 hours；Supernatant is removed in centrifugation；
Confining liquid closing is added, confining liquid contains 0.1-1% cards pool 35,10-50mmol/ml glycine, reacts 2 hours；
Using fluorescent microsphere diluted to 30-120 times, fluorescent microsphere dilutes formula of liquid：PBS+20% trehaloses or
Person's sucrose+1-3% polysorbas20s；
Capture antibody-fluorescent microballoon mixed liquor sprays to glass fibre membrane with the concentration quantitative of 3.0-8.0ul/cm；
25-60 DEG C dries 1-4 hours, kept dry.
Sample pad is blank glass fiber pad, and the strip for cutting into 1.7cm x 30cm width is spare.
In use, sample pad is added after sample is mixed with dilution.Dilution is the 0.8- containing 1-3% polysorbas20s
1%NaCl.Dilution advantageously reduces interference of other ingredients to detection in sample.Under capillary action, sample liquid is to water suction
The swimming of paper one end, when containing SAA in sample to be tested, SAA forms antigen primary antibody bluk recombination with the capture antibody on fluorescent microsphere
Object, as chromatography acts on, compound moves forward, and reaches anti-with SAA detection antibody (test antibody) formation when detection line
Body-Ag-Ab sandwich complex is gathered at detection line T lines.Unbonded fluorescent microsphere continues to move ahead, and reaches nature controlling line C
When, secondary antibody is combined with the capture antibody on fluorescent microsphere, is assembled at C lines.Entire reaction is completed in 15 minutes.It carries out
Upper machine-read card, T lines and C lines can all generate corresponding fluorescence signal, and the information that fluorescence detector can block according to calibration will be practical
Detection fluorescence signal value brings in preset standard curve the content for calculating SAA into.
The present invention is drawn time-resolved fluoroimmunoassay chromatographic technique by applying the enhanced time-resolved fluorescence microballoon of nanometer
In the detection for entering SAA, binding time resolved fluorometric immune detector effectively raises platform sensitivity, is promoted glimmering to low value
The detection result of optical signal, to realize simple, quick, the inexpensive detection of SAA, and high sensitivity, batch interior, difference between batch
It is small, great convenience is provided for Clinical practice, detection provides method by bed to realize.
Embodiment 1
The test strips of the present embodiment are made by following steps：
Detection line and nature controlling line are coated on NC films.
Wherein, for detection line using SAA detection antibody (Test antibody), a concentration of 0.5mg is anti-higher than this concentration
For body using being used after diluted to concentration range, dilution is PBS+1% trehaloses.
Nature controlling line uses the secondary antibody of anti-SAA capture antibody, dosage 0.5mg.Antibody uses diluted to concentration model
It is used after enclosing, dilution is PBS+1% trehaloses.
The two is drawn on NC films with 1.5ul/cm quantity using quantitative spray film instrument, keeps line-to-line every 0.5cm or more, away from
From NC film edges 0.2mm or more.
25 DEG C dry 5 hours, kept dry.
On the other hand, spraying is connected with the 100nm that SAA captures antibody (Capture antibody) on glass fibre membrane
The enhanced time-resolved fluorescence microballoon of nanometer is to prepare bonding pad.
Specifically, the enhanced time-resolved fluorescence microballoon of nanometer is added to the PBS solution of PH6.0, a concentration of 0.05mol/L
In, and EDC, NHS of final concentration of 10mg/ml is added, react at room temperature 30min；
25000rpm/min centrifuges 10min, and supernatant, PBS is gone to wash 2 times, be dissolved in PBS；
SAA is captured antibody and is washed 3 times using ultra-filtration centrifuge tube, and cleaning solution is 0.05mol/L PB buffer solutions, PH8；
Capture antibody after washing is mixed into coupling with fluorescent microsphere, the weight ratio of control antibody/fluorescent microsphere is 1/20,
Supplementing suitable PB buffer solutions makes PH be equal to 7.0, mixing；
Supernatant is removed in room temperature reaction 4 hours, centrifugation；
Confining liquid closing is added, confining liquid reacts 2 hours containing 0.1% card pool 35,10mmol/ml glycine；
Using fluorescent microsphere diluted to 30 times, fluorescent microsphere dilutes formula of liquid：PBS+20% trehaloses+1% are spat
Temperature 20；
Quantitative (3.0ul/cm), which is sprayed to, is made bonding pad on glass fibre membrane, 25 DEG C dry 4 hours, kept dry.
On the other hand, the strip that the glass fibre membrane of a blank is cut into 1.7cm x 30cm width is spare, as sample
Product pad.
Finally, bottom plate, NC films, blotting paper, bonding pad, sample pad are staggeredly pasted successively, cuts into 3.7mm, be packed into modeling
Material gets stuck.
Join Fig. 1, standard curve is drawn using test strips made from the present embodiment.
Specifically, using the time-resolved fluoroimmunoassay chromatograph test strip of SAA, the SAA standard items for detecting various concentration (take
7 different concentration, respectively 0mg/L, 6.25mg/L, 12.5mg/L, 25mg/L, 50mg/L, 100mg/L, 200mg/L, often
A concentration does 5 Duplicate Samples).After film layer analysis reaction 15 minutes, instrument reads nature controlling line C, detection line T signal, with the sample of detection
Product fluorescent value signal ratio T/C is ordinate, and SAA standard concentrations are abscissa, establish equation and are fitted to standard curve.
The coefficient R of the standard curve it can be seen from Fig. 1 standard curves2It is linear preferable more than 0.99, Ke Yitong
It crosses the standard curve and quantitative analysis is carried out to contained SAA concentration in sample.
Embodiment 2
The test strips of the present embodiment are made by following steps：
Detection line and nature controlling line are coated on NC films.
Wherein, detection line is higher than the antibody of this concentration using SAA detection antibody (Test antibody), a concentration of 2mg
Using being used after diluted to concentration range, dilution is PBS+10% trehaloses+5%NaCl.
Nature controlling line uses the secondary antibody of anti-SAA capture antibody, dosage 2mg.Antibody uses diluted to concentration range
After use, dilution is PBS+10% trehaloses+5%NaCl.
The two is drawn on NC films with 0.8ul/cm quantity using quantitative spray film instrument, keeps line-to-line every 0.5cm or more, away from
From NC film edges 0.2mm or more.
60 DEG C dry 1 hour, kept dry.
On the other hand, spraying is connected with the 300nm that SAA captures antibody (Capture antibody) on glass fibre membrane
The enhanced time-resolved fluorescence microballoon of nanometer is to prepare bonding pad.
Specifically, the enhanced time-resolved fluorescence microballoon of nanometer is added to the PBS solution of PH7.0, a concentration of 0.05mol/L
In, and EDC, NHS of final concentration of 10mg/ml is added, react at room temperature 30min；
15000rpm/min centrifuges 10min, and supernatant, PBS is gone to wash 2 times, be dissolved in PBS；
SAA is captured antibody and is washed 3 times using ultra-filtration centrifuge tube, and cleaning solution is 0.05mol/L PB buffer solutions, PH7；
Capture antibody after washing is mixed into coupling with fluorescent microsphere, the weight ratio of control antibody/fluorescent microsphere is 1/10,
Supplementing suitable PB buffer solutions makes PH be equal to 7.4, mixing；
Supernatant is removed in room temperature reaction 1 hour, centrifugation；
Confining liquid closing is added, confining liquid reacts 2 hours containing 1% card pool 35,50mmol/ml glycine；
Using fluorescent microsphere diluted to 120 times, fluorescent microsphere dilutes formula of liquid：+ 3% tween of PBS+20% sucrose
20；
Quantitative (8.0ul/cm), which is sprayed to, is made bonding pad on glass fibre membrane, 60 DEG C dry 1 hour, kept dry.
On the other hand, the strip that the glass fibre membrane of a blank is cut into 1.7cm x 30cm width is spare, as sample
Product pad.
Finally, bottom plate, NC films, blotting paper, bonding pad, sample pad are staggeredly pasted successively, cuts into 3.7mm, be packed into modeling
Material gets stuck.
Join Fig. 1, standard curve is drawn using test strips made from the present embodiment.
Specifically, using the time-resolved fluoroimmunoassay chromatograph test strip of SAA, the SAA standard items for detecting various concentration (take
7 different concentration, respectively 0mg/L, 6.25mg/L, 12.5mg/L, 25mg/L, 50mg/L, 100mg/L, 200mg/L, often
A concentration does 5 Duplicate Samples).After film layer analysis reaction 15 minutes, instrument reads nature controlling line C, detection line T signal, with the sample of detection
Product fluorescent value signal ratio T/C is ordinate, and SAA standard concentrations are abscissa, establish equation and are fitted to standard curve.
The coefficient R of the standard curve it can be seen from Fig. 1 standard curves2It is linear preferable more than 0.99, Ke Yitong
It crosses the standard curve and quantitative analysis is carried out to contained SAA concentration in sample.
Embodiment 3
30 patients serum's samples are randomly selected, use Siemens BII scattering turbidimetry instrument test datas as reference point, with this
The numerical value for test corresponding sample under invention test strips room temperature, with statistical analysis system coherence.
The application method of test strips of the present invention is：It is added in the sample application zone of the fluorescence immune chromatography test paper bar of SAA and waits for test sample
Product, film layer analysis reaction 15 minutes；Fluorescence detection device is opened, and detector bar and calibration card are included and include card into fluorescence detection device
Mouthful, instrument is run, calculates the SAA concentration in sample to be tested automatically by corresponding analysis software.
Join Fig. 3 and Fig. 4, carries out contrast experiment respectively using test strips made from above-described embodiment 1 and embodiment 2, as a result
It has been shown that, the testing result and Siemens SAA testing result othernesses of test strips of the present invention are small, linearly dependent coefficient R2It respectively reaches
0.9785 and 0.9835.
It should be appreciated that although this specification is described in terms of embodiments, but not each embodiment only includes one
A independent technical solution, this description of the specification is merely for the sake of clarity, and those skilled in the art should will say
As a whole, the technical solution in each embodiment may also be suitably combined to form those skilled in the art can for bright book
With the other embodiment of understanding.
The series of detailed descriptions listed above only for the present invention feasible embodiment specifically
Bright, they are all without departing from equivalent implementations made by technical spirit of the present invention not to limit the scope of the invention
Or change should all be included in the protection scope of the present invention.
Claims (10)
1. a kind of preparation method of the time-resolved fluoroimmunoassay chromatograph test strip of quantitatively detection serum amyloid A protein, special
Sign is, the method includes,
Nitrocellulose filter is coated with detection line and nature controlling line, and the detection line detects antibody using serum amyloid A protein, described
Nature controlling line captures the secondary antibody of antibody using antiserum amyloid A；
Prepare bonding pad comprising：
In PBS containing the enhanced time-resolved fluorescence microballoon of nanometer, the EDC and NHS of final concentration of 10mg/ml is added, centrifuges,
Precipitation is dissolved in after PBS is washed in PBS；
Serum amyloid A protein captures antibody and uses antibody cleaning solution centrifuge washing；
Serum amyloid A protein capture antibody after washing is mixed with the PBS dissolved with the fluorescent microsphere, so that serum forms sediment
Powder sample albumin A captures antibody and is coupled with fluorescent microsphere；
Mixed liquor after coupling is sprayed into carrier；
Wherein, PBS PH6-7, concentration 0.05mol/L.
2. preparation method according to claim 1, which is characterized in that a diameter of 100nm-300nm of fluorescent microsphere.
3. preparation method according to claim 1, which is characterized in that serum amyloid A protein capture antibody with it is glimmering
The weight ratio of light microballoon mixing is 1/10-1/20.
4. preparation method according to claim 1, which is characterized in that serum amyloid A protein capture antibody with it is described glimmering
It after the coupling of light microballoon, is closed through confining liquid, then sprays to carrier, it is 0.1-1%'s that the confining liquid, which contains mass percent,
Block the glycine of 35 and a concentration of 10-50mmol/ml of pool.
5. preparation method according to claim 1, which is characterized in that the antibody cleaning solution is the 0.05mol/L of PH7-8
PB buffer solutions.
6. preparation method according to claim 1, which is characterized in that the carrier is glass fibre membrane.
7. preparation method according to claim 1, which is characterized in that after antibody is mixed with fluorescent microsphere, further include
Step：Serum amyloid A protein capture antibody and the pH value of fluorescent microsphere mixed liquor are adjusted to 7.2 ± 0.2.
8. preparation method according to claim 1, which is characterized in that the serum amyloid A protein of each detection line
Detection antibody dosage is 0.5-2mg.
9. preparation method according to claim 1, which is characterized in that the antiserum amyloid protein of each nature controlling line
A captures the secondary antibody of antibody, dosage 0.5-2mg.
10. it is a kind of quantitatively detect serum amyloid A protein time-resolved fluoroimmunoassay chromatograph test strip, by bottom plate, sample pad,
Bonding pad, nitrocellulose filter and blotting paper composition, which is characterized in that the test strips are any described by claim 1-9
Method is made.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810298348.1A CN108680751A (en) | 2018-04-04 | 2018-04-04 | Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810298348.1A CN108680751A (en) | 2018-04-04 | 2018-04-04 | Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108680751A true CN108680751A (en) | 2018-10-19 |
Family
ID=63799792
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810298348.1A Pending CN108680751A (en) | 2018-04-04 | 2018-04-04 | Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108680751A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109613256A (en) * | 2018-11-21 | 2019-04-12 | 杭州康知生物科技有限公司 | It urinates four kinds of traces of albumin and quantifies joint inspection fluorescence immune chromatography kit and preparation method |
| CN110618266A (en) * | 2019-09-26 | 2019-12-27 | 河北省科学院生物研究所 | Kit for detecting mastitis of dairy cow and use method thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5262303A (en) * | 1989-10-13 | 1993-11-16 | Trustees Of Boston University | Ligand/anti-ligand assays for adherent proteins |
| JPH08178921A (en) * | 1994-12-21 | 1996-07-12 | Cosmo Sogo Kenkyusho:Kk | Immunoassay of serum amyloid a |
| WO1997004317A1 (en) * | 1995-07-21 | 1997-02-06 | The Provost, Fellows And Scholars Of The College Of The Holy And Undivided Trinity Of Queen Elizabeth Near Dublin | Method for the quantitative measurement of human acute phase serum amyloid a protein; recombinant protein; specific antibody |
| CN105092861A (en) * | 2015-09-14 | 2015-11-25 | 广州市微米生物科技有限公司 | Immunofluorescence chromatography test paper for CRP (C-reaction protein)/SAA (Serum amyloid A protein) quantitative combined detection and preparation method of immunofluorescence chromatography test paper |
| CN106706926A (en) * | 2016-12-16 | 2017-05-24 | 威海纽普生物技术有限公司 | Serum amyloid A testing kit and manufacturing method |
-
2018
- 2018-04-04 CN CN201810298348.1A patent/CN108680751A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5262303A (en) * | 1989-10-13 | 1993-11-16 | Trustees Of Boston University | Ligand/anti-ligand assays for adherent proteins |
| JPH08178921A (en) * | 1994-12-21 | 1996-07-12 | Cosmo Sogo Kenkyusho:Kk | Immunoassay of serum amyloid a |
| WO1997004317A1 (en) * | 1995-07-21 | 1997-02-06 | The Provost, Fellows And Scholars Of The College Of The Holy And Undivided Trinity Of Queen Elizabeth Near Dublin | Method for the quantitative measurement of human acute phase serum amyloid a protein; recombinant protein; specific antibody |
| CN105092861A (en) * | 2015-09-14 | 2015-11-25 | 广州市微米生物科技有限公司 | Immunofluorescence chromatography test paper for CRP (C-reaction protein)/SAA (Serum amyloid A protein) quantitative combined detection and preparation method of immunofluorescence chromatography test paper |
| CN106706926A (en) * | 2016-12-16 | 2017-05-24 | 威海纽普生物技术有限公司 | Serum amyloid A testing kit and manufacturing method |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109613256A (en) * | 2018-11-21 | 2019-04-12 | 杭州康知生物科技有限公司 | It urinates four kinds of traces of albumin and quantifies joint inspection fluorescence immune chromatography kit and preparation method |
| CN110618266A (en) * | 2019-09-26 | 2019-12-27 | 河北省科学院生物研究所 | Kit for detecting mastitis of dairy cow and use method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN108279309B (en) | Detection test strip and detection method for PLA2R antibody | |
| US4054646A (en) | Method and apparatus for detection of antibodies and antigens | |
| CN102628867A (en) | Double antibody latex enhanced retinol binding protein detection kit | |
| CN102590526A (en) | Beta 2-microglobulin detection kit | |
| CN106370861B (en) | A kind of c reactive protein saliva test strip and preparation method thereof | |
| CN105403693A (en) | Preparation method of magnetic particle chemiluminescence reagent | |
| CN107870234A (en) | A kind of test strips for detecting thyroglobulin concentration and preparation method thereof | |
| CN101226195A (en) | Kit for detecting knubble type M2 pyruvate kinase and preparation method thereof | |
| CN105717308A (en) | Immunochromatography kit for fast and quantitatively detecting fecal lactoferrin | |
| CN101650368A (en) | Method for testing zearalenone toxin by using colloidal gold immunochromatography assay | |
| CN108680751A (en) | Quantitatively detect the time-resolved fluoroimmunoassay chromatograph test strip and preparation method thereof of serum amyloid A protein | |
| JPH09508975A (en) | Analytical cuvette, quantification method and diagnostic test kit used for quantification of components to be analyzed present in whole blood sample | |
| CN106771239A (en) | Serum amyloid A protein/Procalcitonin/C reactive proteins are three-in-one to determine kit and preparation method | |
| CN102539786B (en) | Microscale urinary albumin colloidal gold detection kit and preparation technology thereof | |
| CN206038696U (en) | Ration calprotectin detects immunochromatographic test strip | |
| CN108061727A (en) | A kind of thyroglobulin quantitative testing test paper item and preparation method thereof | |
| EP2982987A1 (en) | Lateral flow immunoassays for the detection of antibodies against biological drugs | |
| CN101046479B (en) | Process of preparing human serum base matter containing no target protein | |
| CN109459570A (en) | The colloid gold immune test paper preparation method of lead ion in a kind of quick detection blood of human body | |
| JP2009258095A (en) | Method for manufacturing strip for chromatographic assay | |
| CN103267842B (en) | Detect the immune colloidal gold method of the illegal interpolation Diclofenac of Chinese patent drug | |
| CN202916284U (en) | NGAL (neutrophil gelatinase associated lipocalin) whole blood detecting device | |
| CN206038688U (en) | Immunity chromatography detection test strip's fluorescent quantitation spectral detection system | |
| CA2116687A1 (en) | Initial rate photometric method for immunoassay | |
| CN202837303U (en) | Neutrophil gelatinase associated lipocalin (NGAL) fluorescence immunochromatography quantitative detection test strip |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20181019 |