CN109724929A - A kind of detection method of anti-bad citrulling peptide antibody - Google Patents
A kind of detection method of anti-bad citrulling peptide antibody Download PDFInfo
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- CN109724929A CN109724929A CN201811604492.XA CN201811604492A CN109724929A CN 109724929 A CN109724929 A CN 109724929A CN 201811604492 A CN201811604492 A CN 201811604492A CN 109724929 A CN109724929 A CN 109724929A
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- citrulling peptide
- citrulling
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Abstract
The invention discloses a kind of detection methods of anti-bad citrulling peptide antibody, including anti-bad citrulling peptide antigen, anti-bad citrulling peptide antibody standard items, preservative and stabilizer；The anti-bad citrulling peptide antigen obtains anti-bad citrulling peptide antigen latex by processing.Anti- its surface of bad citrulling peptide antigen emulsion reagent, which is made, in detection method provided by the invention can adsorb more multispecific antibody, can further promote the sensitivity of detection.
Description
Technical field
The present invention relates to field of biotechnology, more particularly to a kind of detection method of anti-bad citrulling peptide antibody.
Background technique
Rheumatoid arthritis is a kind of systemic autoimmune disease characterized by chornic arthritis, and the disease is for example too late
When diagnoses and treatment, that is, may occur in which the aggressivity lesion of cartilage and bone, joint structure caused to destroy, be eventually developed to for joint it is abnormal
It is shape, tetanic, lead to different degrees of deformity, therefore early diagnose most important.In recent years, research shows that antiCCP antibody is early
Phase diagnoses the good sign object of RA, diagnoses the sensitivity 41%~67.6% of RA, specificity 70.9% one 89%.One kind is provided
Convenient, fast, efficient CCP detection method is technical problem urgently to be solved in the prior art.
Summary of the invention
In view of the above-mentioned problems, the present invention provides a kind of detection method of anti-bad citrulling peptide antibody, to solve the prior art
Present in technical problem.
To realize above-mentioned technical purpose and the technique effect, the invention is realized by the following technical scheme:
A kind of detection method of anti-bad citrulling peptide antibody, including anti-bad citrulling peptide antigen, anti-bad citrulling peptide antibody standard
Product, preservative and stabilizer；The anti-bad citrulling peptide antigen is handled by following steps:
A) the MES solution 10ml of 20mmol/L is taken, the magnesium chloride solution 0.2ml of 0.02mol/L is added thereto, in 6-80C water
10 minutes are kept the temperature in bath；
B) anti-bad citrulling peptide antigen is added into the resulting solution of step a, stands 5 minutes；
C) purification buffer, ultrasound 3 minutes are added into the polystyrene colloidal milk solution that partial size is 100-350nm；
D) step b acquired solution is added in step c acquired solution and forms mixed liquor, pH value is added in Xiang Shangshu mixed liquor is
The confining liquid of 7.0-7.8 is in 25-3002 hours or more are stood in C environment；
E) by taking precipitate after the centrifugation of step d acquired solution；
F) PBS buffer solution that PH is 6.5 of the sediment in step e is filtered at least twice, obtains anti-bad citrulling peptide antigen
Substance obtained above is loaded into kit and forms detection kit by latex.
Preferably, the purification buffer solution is Tris- phosphate buffer, and the PH of buffer is 6.2-7.3.
Preferably, the calcium chloride solution 0.5ml containing 0.05mol/L in every 10ml purification buffer solution.
Preferably, the purification buffer is put into 50030 minutes or more are kept the temperature in C water-bath, is then naturally cooled to again
250C。
Preferably, the anti-bad citrulling peptide antibody standard items are the anti-bad citrulling peptide antibody of 96-100U/ml, 3-15mg/
The stabilizer of the PBS buffer solution of ml, 20-60ml.
Preferably, the stabilizer is selected from one of surfactant, antioxidant, suspending agent or a variety of.
Preferably, the preservative is selected from sodium azide or gentamicin.
The beneficial effects of the present invention are:
Since CCP is the small molecule for containing only 21 amino acid, direct coated is not easy in solid phase carrier, is examined for antiCCP antibody
Survey will lead to Lower result often.Anti- bad its surface energy of citrulling peptide antigen emulsion reagent is made in detection method provided by the invention
More multispecific antibody is adsorbed, the sensitivity of detection can be further promoted.
Detailed description of the invention
Fig. 1 is CCP reference standard curve.
Specific embodiment
Technical solution of the present invention is described in further detail with specific embodiment with reference to the accompanying drawing, so that ability
The technical staff in domain can better understand the present invention and can be practiced, but illustrated embodiment is not as to limit of the invention
It is fixed.
Embodiment 1
The detection method of anti-bad citrulling peptide antibody in the present embodiment, including anti-bad citrulling peptide antigen, anti-bad citrulling peptide
Antibody standard substance, preservative and stabilizer；
The anti-bad citrulling peptide antigen is handled by following steps:
A) the MES solution 10ml of 20mmol/L is taken, the magnesium chloride solution 0.2ml of 0.02mol/L is added thereto, in 6-80C water
10 minutes are kept the temperature in bath；
B) anti-bad citrulling peptide antigen is added into the resulting solution of step a, stands 5 minutes；
C) purification buffer, ultrasound 3 minutes are added into the polystyrene colloidal milk solution that partial size is 100-350nm；
D) step b acquired solution is added in step c acquired solution and forms mixed liquor, pH value is added in Xiang Shangshu mixed liquor is
The confining liquid of 7.0-7.8 is in 25-3002 hours or more are stood in C environment；
E) by taking precipitate after the centrifugation of step d acquired solution；
F) PBS buffer solution that PH is 6.5 of the sediment in step e is filtered at least twice, obtains anti-bad citrulling peptide antigen
Latex.
Above-mentioned purification buffer solution is Tris- phosphate buffer, and the PH of buffer is 6.2-7.3.Every 10ml purification is slow
Rush the calcium chloride solution 0.5ml in solution containing 0.05mol/L.The purification buffer is put into 50030 minutes are kept the temperature in C water-bath
More than, 25 are then naturally cooled to again0C。
Anti- bad citrulling peptide antibody standard items are the PBS buffering of the anti-bad citrulling peptide antibody of 96-100U/ml, 3-15mg/ml
The stabilizer of liquid, 20-60ml.
Embodiment 2
The kit of the measuring method of anti-bad citrulling peptide antibody detection reagent, this method production is suitable for the biochemistry of a variety of brands
Analyzer, measuring method are as follows:
1 determination condition: temperature 370C, dominant wavelength: 550nm, commplementary wave length 800nm, sample size 5uL, buffer solution 180ml, resist
Bad citrulling peptide antigen latex 50uL, classifying type: Two point end assay
The absorbance for first measuring blank after 5uL sample is added in 180ml buffer, preincubate 3 minutes, measures Blank absorbance
The anti-bad citrulling peptide antigen latex of 50uL is added after degree, is incubated for 3 minutes, measures back suction luminosity.
The above is only a preferred embodiment of the present invention, is not intended to limit the scope of the invention, all to utilize this hair
Equivalent structure made by bright specification and accompanying drawing content perhaps equivalent process transformation or be directly or indirectly used in other correlation
Technical field, be included within the scope of the present invention.
Claims (7)
1. a kind of detection method of anti-bad citrulling peptide antibody characterized by comprising anti-bad citrulling peptide antigen, anti-bad melon
Propylhomoserin peptide antibody standard items, preservative and stabilizer；
The anti-bad citrulling peptide antigen is handled by following steps:
The MES solution 10ml of 20mmol/L is taken, the magnesium chloride solution 0.2ml of 0.02mol/L is added thereto, in 6-80In C water-bath
Heat preservation 10 minutes；
Anti- bad citrulling peptide antigen is added into the resulting solution of step a, stands 5 minutes；
Purification buffer, ultrasound 3 minutes are added into the polystyrene colloidal milk solution that partial size is 100-350nm；
Step b acquired solution is added in step c acquired solution and forms mixed liquor, it is 7.0- that pH value is added in Xiang Shangshu mixed liquor
7.8 confining liquid is in 25-3002 hours or more are stood in C environment；
By taking precipitate after the centrifugation of step d acquired solution；
Sediment in the step e PBS buffer solution that PH is 6.5 is filtered at least twice, anti-bad citrulling peptide antigen glue is obtained
Cream.
2. the detection method of anti-bad citrulling peptide antibody according to claim 1 is it is characterized in that, purification buffering is molten
Liquid is Tris- phosphate buffer, and the PH of buffer is 6.2-7.3.
3. the detection method of anti-bad citrulling peptide antibody according to claim 2 is it is characterized in that, every 10ml purification buffering
Calcium chloride solution 0.5ml in solution containing 0.05mol/L.
4. the detection method of anti-bad citrulling peptide antibody according to claim 3 is it is characterized in that, the purification buffer
It is put into 50030 minutes or more are kept the temperature in C water-bath, then naturally cools to 25 again0C。
5. the detection method of anti-bad citrulling peptide antibody according to claim 1 is it is characterized in that, the anti-bad citrulling
Peptide antibody standard items are the anti-bad citrulling peptide antibody of 96-100U/ml, the PBS buffer solution of 3-15mg/ml, the stabilization of 20-60ml
Agent.
6. the detection method of anti-bad citrulling peptide antibody according to claim 1 is it is characterized in that, the stabilizer is selected from
One of surfactant, antioxidant, suspending agent are a variety of.
7. the detection method of anti-bad citrulling peptide antibody according to claim 1 is it is characterized in that, the preservative is selected from
Sodium azide or gentamicin.
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|---|---|---|---|
| CN201811604492.XA CN109724929A (en) | 2018-12-26 | 2018-12-26 | A kind of detection method of anti-bad citrulling peptide antibody |
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| CN201811604492.XA CN109724929A (en) | 2018-12-26 | 2018-12-26 | A kind of detection method of anti-bad citrulling peptide antibody |
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102507918A (en) * | 2011-11-09 | 2012-06-20 | 四川省新成生物科技有限责任公司 | Kit for determining anti-cyclic citrullinated peptide (Anti-CCP) antibody |
| CN104198725A (en) * | 2014-08-14 | 2014-12-10 | 上海睿康生物科技有限公司 | Anti-cyclic citrullinated peptide (CCP) antibody detection kit |
| CN104764888A (en) * | 2015-05-04 | 2015-07-08 | 潍坊市康华生物技术有限公司 | Anti-cyclic citrullinated peptide antibody detection reagent kit |
| CN106153946A (en) * | 2016-06-22 | 2016-11-23 | 浙江达美生物技术有限公司 | A kind of mensuration reagent of anti-cyclic citrullinated peptide and preparation method thereof |
| CN107942067A (en) * | 2017-11-15 | 2018-04-20 | 浙江夸克生物科技有限公司 | Cyclic citrullinated peptid assay kit |
| CN108614122A (en) * | 2018-07-27 | 2018-10-02 | 金华市强盛生物科技有限公司 | A kind of cyclic citrullinated peptid assay kit |
-
2018
- 2018-12-26 CN CN201811604492.XA patent/CN109724929A/en not_active Withdrawn
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102507918A (en) * | 2011-11-09 | 2012-06-20 | 四川省新成生物科技有限责任公司 | Kit for determining anti-cyclic citrullinated peptide (Anti-CCP) antibody |
| CN104198725A (en) * | 2014-08-14 | 2014-12-10 | 上海睿康生物科技有限公司 | Anti-cyclic citrullinated peptide (CCP) antibody detection kit |
| CN104764888A (en) * | 2015-05-04 | 2015-07-08 | 潍坊市康华生物技术有限公司 | Anti-cyclic citrullinated peptide antibody detection reagent kit |
| CN106153946A (en) * | 2016-06-22 | 2016-11-23 | 浙江达美生物技术有限公司 | A kind of mensuration reagent of anti-cyclic citrullinated peptide and preparation method thereof |
| CN107942067A (en) * | 2017-11-15 | 2018-04-20 | 浙江夸克生物科技有限公司 | Cyclic citrullinated peptid assay kit |
| CN108614122A (en) * | 2018-07-27 | 2018-10-02 | 金华市强盛生物科技有限公司 | A kind of cyclic citrullinated peptid assay kit |
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Application publication date: 20190507 |