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Assay method for antibodies against cyclic citrullinated peptide

An antibody and main body technology, applied in the field of detection of anti-cyclic citrullinated peptide antibodies, can solve the problems of low reliability and difficult detection, etc.

Inactive Publication Date: 2017-12-01
AXIS SHIELD DIAGNOSTICS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It is also difficult to automate such detection, and when automated, the reliability is relatively low

Method used

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  • Assay method for antibodies against cyclic citrullinated peptide
  • Assay method for antibodies against cyclic citrullinated peptide
  • Assay method for antibodies against cyclic citrullinated peptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0085] Turbidity detection of anti-CCP antibody

[0086] (a) Preparation of avidin-coated nanoparticles

[0087] 600 μl of 4.2% w / v chloromethyl-activated nanoparticles (diameter 44 nm) (purchased from Interfacial Dynamic Corporation, USA) were subjected to water dialysis with a membrane with a pore size of 300,000 Da.

[0088] 0.5 ml of borate (10 mM) and sodium chloride (15 mM) solution (pH 9.0) was added and mixed. 10 mg of avidin dissolved in 0.5 ml of a 10 mM borate and 15 mM NaCl solution (purchased from Pierce Chemical Company) at pH 9.0 was added and the mixture was stirred at room temperature for 24 hours. Then 40 μl of a glycine solution (2M, pH 9.0) was added and the mixture was stirred for a further 4 hours at room temperature.

[0089]The microparticles were then diluted to a volume of 100 ml and dialyzed, first in 500 ml of 10 mM borate and 15 mM sodium chloride solution (pH 9.0), then in 25 mM Tris, 150 mM sodium chloride and 0.01% Tween 20 solution ( pH 7.4)...

Embodiment 2

[0094] Example 2: Turbidity Detection of Alternative Anti-CCP Antibodies

[0095] (a) Preparation of CCP peptide-coated nanoparticles

[0096] 1 ml of 4.2% w / v chloromethyl-activated nanoparticles (diameter 44 nm) (purchased from Interfacial Dynamic Corporation, USA) was subjected to water dialysis using a membrane with a pore size of 300,000 Da.

[0097] Then 0.5 ml of a 10 mM borate and 15 mM sodium chloride solution (pH 9.0) was added. 1.5 μg of purified CCP peptide (eg, affinity purified CCP peptide) was dialyzed against 10 mM borate and 15 mM sodium chloride solution (pH 9.0).

[0098] After the addition of nanoparticles to the purified CCP peptide, the mixture was stirred at room temperature for 24 hours. Then 40 [mu]l of glycine solution (2M, pH 9.0) was added and the mixture was stirred for a further 4 hours at room temperature.

[0099] The microparticles were then diluted to a total volume of 100 ml and dialyzed against 1000 ml of a 10 mM borate and 15 mM sodium c...

Embodiment 3

[0106] Example 3: Turbidity detection of anti-CCP antibody

[0107] (a) Preparation of streptavidin-coated nanoparticles

[0108] 600 μl of 4.2% w / v chloromethyl-activated nanoparticles (diameter 67 nm) (purchased from Interfacial Dynamic Corporation, USA) were subjected to water dialysis with a membrane with a pore size of 300,000 Da.

[0109] 0.5 ml of phosphate (10 mM) and sodium chloride (150 mM) buffer (pH 7.4) and 10 mg of streptavidin dissolved in 0.5 ml of 10 mM phosphate and 150 mM NaCl buffer (pH 7.4) (available from Pierce Chemical Company) were added together and the resulting mixture was stirred at room temperature for 24 hours. Then 40 [mu]l of glycine solution (2M, pH 9.0) was added and the resulting mixture was stirred for a further 4 hours at room temperature.

[0110] The microparticles were then diluted to a volume of 100 ml and dialyzed, first in 500 ml of 10 mM borate and 15 mM sodium chloride solution (pH 9.0), then in 25 mM Tris, 150 mM sodium chloride...

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Abstract

The present invention relates to method for assaying anti-cyclic citrullinated peptide antibodies in a clinical sample, said method comprising contacting said sample with at least one homogeneous reagent comprising at least one specific binder for anti-CCP antibodies, whereby to form a solution or suspension of an anti-CCP-binding partner complex in a homogeneous sample mixture and detecting the presence or level of said anti-CCP-binding partner complex in a homogeneous liquid-phase. The invention also relates to a method for the assessment of the existence of; risk of; potential for; or propensity to RA in a subject, said method comprising assaying for anti-CCP in a body sample from said subject in a homogeneous assay according to the invention, whereby to determine the level of anti-CCP antibodies in said sample, and correlating the thus-determined level with the existence of; risk of; potential for; or propensity to RA in said subject.

Description

[0001] This application is a divisional application of an invention patent application with an application date of February 20, 2009, an application number of 200980113814.3, and an invention title of "Detection method for anti-cyclic citrullinated peptide antibody". technical field [0002] The present invention relates to in vitro assays for detecting the presence, potential or predisposition of rheumatoid arthritis (RA) in a subject. In particular, the present invention relates to automated tests, and in particular to tests for the assessment of rheumatoid arthritis (RA) comprising the detection of endogenous antibodies (anti-CCP antibodies) specific for cyclic citrullinated peptides in a subject. presence or level. The invention also relates to methods of detecting the presence or level of endogenous antibodies specific to cyclic citrullinated peptides (anti-CCP antibodies) in a subject. Background technique [0003] Rheumatoid arthritis (RA) is a common, systemic autoi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/564
CPCG01N33/564G01N2800/102
Inventor 肯·米尔恩戴维·普里查德埃尔林·松德热哈根
Owner AXIS SHIELD DIAGNOSTICS
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