WO2002063302A1 - Technique permettant de reduire l'interference de l'heparine dans des essais diagnostic de la troponine cardiaque - Google Patents

Technique permettant de reduire l'interference de l'heparine dans des essais diagnostic de la troponine cardiaque Download PDF

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Publication number
WO2002063302A1
WO2002063302A1 PCT/US2002/003713 US0203713W WO02063302A1 WO 2002063302 A1 WO2002063302 A1 WO 2002063302A1 US 0203713 W US0203713 W US 0203713W WO 02063302 A1 WO02063302 A1 WO 02063302A1
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WO
WIPO (PCT)
Prior art keywords
heparin
antagonist
antibody
moiety
troponin
Prior art date
Application number
PCT/US2002/003713
Other languages
English (en)
Inventor
Lance A. Liotta
Alan R. Day
Heather Schessler
Tabitha W. Harlacher
Original Assignee
Immunomatrix Inc.
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Immunomatrix Inc. filed Critical Immunomatrix Inc.
Publication of WO2002063302A1 publication Critical patent/WO2002063302A1/fr

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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
    • G01N33/6887—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids from muscle, cartilage or connective tissue
    • 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
    • G01N2333/4701—Details
    • G01N2333/4712—Muscle proteins, e.g. myosin, actin, protein

Definitions

  • the present invention relates to assays for diagnostic testing. More particularly, the invention is directed to processes for reducing anticoagulant interferences in diagnostic tests, such as diagnostic immunoassays for cardiac Troponin
  • Cardiac Troponin is a complex of three proteins of the thin filament of muscle. It is a muscle protein which functions to regulate muscle contraction.
  • Cardiac Troponin I measurement has become the method of choice in detection of cardiac muscle damage because it's structure in heart muscle is unique muscle found in other parts of the body. The measurement has been facilitated by the generation of antibodies directed to the cardiac specific regions of Troponin I and incorporating these into commercially available assays. The major issue with these assays is that there is a lack of both standardization and correlation among methods. (Wu, A.H.B. Laboratory and Near Patient Testing for Cardiac Markers. J. Clin Ligand Assay, 22: 32-27 (1999)).
  • heparin binds to cardiac Troponin I (cTNI) via a charge- charge interaction.
  • cTNI cardiac Troponin I
  • Heparin is frequently found in blood derived as a compound in samples because it is used as an anti-coagulant for sample collection or it is being used as a treatment in cardiac patients.
  • the interaction of heparin with both free and complexed Troponin I can further complicate the assay results. It is reported by one manufacturer that the bias seen with heparin samples is 30% lower than in serum or other plasma samples (Dade Behring Opus plus Troponin I instructions for use). Consequently, it would be desirable if such interference could be overcome in order to provide a more accurate result. It would also be desirable if the correlation of different assay methods could be improved with the elimination of an interfering factor.
  • the present invention is directed to a process for reducing heparin anticoagulant interference binding in an antibody-antigen diagnostic assay, such as a diagnostic assay for cardiac Troponin I.
  • the process utilizes a moiety, such as a highly charged peptide, which competes with heparin at the antibody-antigen binding site.
  • the moiety is a heparin antagonist such as a protamine salt or hexadimethrine bromide (polybrene).
  • the invention relates to a process for increasing the sensitivity of an antibody-antigen diagnostic assay for cardiac Troponin I.
  • the process utilizes a moiety such as a highly charged peptide which competes with heparin at the antibody-antigen binding site.
  • the moiety is a charged peptide with a sequence similar to the native sequence but sufficient different to allow for a specific antibody-antigen reaction.
  • the moiety is a heparin antagonist, and most preferably, the moiety is a protamine salt such as protamine sulfate or polybrene.
  • the present invention is directed to a quantitative strip base immunoassay such as a quantitative strip base luminescent immunoassay to be used in the point of care environment.
  • the immunoassay is useful for the testing for indications of acute myocardial infarction.
  • the immunoassay can be utilized in conjunction with a moiety which competes with heparin at the antibody-antigen binding site or the moiety can be included in strip based immunoassay, such as with a solubilized substrate, etc.
  • the invention accordingly comprises several components or steps and the relation of one or more of such components or steps with respect to each of the others and the article possessing the features, properties, and the relation of elements exemplified in the following detailed disclosure.
  • BRIEF DESCRIPTION OF THE DRAWINGS [00011]
  • the invention may take form in various components and arrangements of components and in various steps and arrangements of steps.
  • the drawings are only for purposes of illustrating preferred embodiments and are not to be construed as limiting the invention.
  • Fig. 1 is a listing of the skeletal and cardiac amino acid sequences for human Troponin I;
  • Fig. 2 is a chart illustrating the kinetics of cardiac Troponin I in three human matrices
  • Fig. 3 is a graph showing the effect of protamine against heparine using an immunoassay for cardiac Troponin I;
  • Fig. 4 is a chart demonstrating the effect produced by the heparin antagonists, polybrene and protamine, on an immunoassay for cardiac Troponin I;
  • Fig. 5 is an illustration of quantitative strip based luminescent immunoassay containing, or utilized in conjunction with, a heparin antagonist.
  • SeraCare, San Diego CA were compared in spike and recovery experiments using human cTNI (Hytest, Finland) preparations at estimated values of 25, 50 and 75 ng/ml concentrations.
  • Recovery in a prototype cTNI assay (Immunomatrix, Gaithersburg, MD) showed between 50 and 91% lower values in the heparinized plasma than the fresh frozen plasma.
  • an examination of the kinetics of binding was performed using heparinized plasma (SeraCare, San Diego), heparinized plasma with the addition of 2mg/ml protamine sulfate salt (Sigma, St. Louis MO) and serum (in house pool) all spiked with 20ng/ml of human cTNI (Hytest, Finland).
  • heparin antagonists There are other heparin antagonists available commercially.
  • polybrene hexadimethrine bromide
  • cTNI immunomatrix prototype cTNI assay as described in example 3.
  • samples prepared from heparinized plasma with either 2 mg/ml protamine or 1 mg/ml polybrene and spiked with levels of cTNI at 0, a low concentration (1 ng/ml) or a high concentration (50 ng/ml) an evaluation of the effect of these two heparin antagonists was made.

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Hematology (AREA)
  • Immunology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Cell Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Peptides Or Proteins (AREA)

Abstract

La présente invention concerne un processus permettant de réduire l'interférence de l'anticoagulant héparine se liant dans l'essai diagnostic anticorps-antigène, tel qu'un essai diagnostic de la troponine cardiaque. Ce processus utilise une fraction, telle qu'un peptide hautement chargé, qui est en concurrence avec l'éparine au niveau du site de liaison anticorps-antigène. Cette fraction est de préférence un antagoniste d'héparine tel qu'un sel de protamine ou qu'un bromure hexadiméthrine (polybrène).
PCT/US2002/003713 2001-02-07 2002-02-07 Technique permettant de reduire l'interference de l'heparine dans des essais diagnostic de la troponine cardiaque WO2002063302A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US26689201P 2001-02-07 2001-02-07
US60/266,892 2001-02-07

Publications (1)

Publication Number Publication Date
WO2002063302A1 true WO2002063302A1 (fr) 2002-08-15

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2002/003713 WO2002063302A1 (fr) 2001-02-07 2002-02-07 Technique permettant de reduire l'interference de l'heparine dans des essais diagnostic de la troponine cardiaque

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WO (1) WO2002063302A1 (fr)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108254579A (zh) * 2018-01-31 2018-07-06 济南景正医疗器械有限公司 制备一种抗干扰能力强的高密度脂蛋白胆固醇检测试剂盒
JPWO2017179611A1 (ja) * 2016-04-13 2019-02-21 株式会社Ｌｓｉメディエンス 硫酸化多糖類を用いた免疫学的測定法
US10670611B2 (en) 2014-09-26 2020-06-02 Somalogic, Inc. Cardiovascular risk event prediction and uses thereof

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5320812A (en) * 1993-10-04 1994-06-14 Becton, Dickinson And Company Clot activating polyelectrolyte complex and blood collection assembly containing same
US5677133A (en) * 1993-02-17 1997-10-14 Cardiovascular Diagnostics, Inc. Dry chemistry cascade immunoassay and affinity assay
WO1999040442A1 (fr) * 1998-02-05 1999-08-12 Bio-Rad Pasteur Dosage des troponines sans interferences dues a l'heparine
US6174686B1 (en) * 1995-04-18 2001-01-16 Biosite Diagnostics, Inc. Methods for the assay of troponin I and T and complexes of troponin I and T and selection of antibodies for use in immunoassays

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5677133A (en) * 1993-02-17 1997-10-14 Cardiovascular Diagnostics, Inc. Dry chemistry cascade immunoassay and affinity assay
US5320812A (en) * 1993-10-04 1994-06-14 Becton, Dickinson And Company Clot activating polyelectrolyte complex and blood collection assembly containing same
US6174686B1 (en) * 1995-04-18 2001-01-16 Biosite Diagnostics, Inc. Methods for the assay of troponin I and T and complexes of troponin I and T and selection of antibodies for use in immunoassays
WO1999040442A1 (fr) * 1998-02-05 1999-08-12 Bio-Rad Pasteur Dosage des troponines sans interferences dues a l'heparine

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US10670611B2 (en) 2014-09-26 2020-06-02 Somalogic, Inc. Cardiovascular risk event prediction and uses thereof
JPWO2017179611A1 (ja) * 2016-04-13 2019-02-21 株式会社Ｌｓｉメディエンス 硫酸化多糖類を用いた免疫学的測定法
EP3444612A4 (fr) * 2016-04-13 2019-12-11 LSI Medience Corporation Dosage immunologique utlisant un polysaccharide sulfaté
US11422128B2 (en) 2016-04-13 2022-08-23 Lsi Medience Corporation Immunoassay employing sulfated polysaccharide
CN108254579A (zh) * 2018-01-31 2018-07-06 济南景正医疗器械有限公司 制备一种抗干扰能力强的高密度脂蛋白胆固醇检测试剂盒

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