WO2007117155A1 - Essai immunologique en phase solide - Google Patents

Essai immunologique en phase solide Download PDF

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Publication number
WO2007117155A1
WO2007117155A1 PCT/NZ2007/000044 NZ2007000044W WO2007117155A1 WO 2007117155 A1 WO2007117155 A1 WO 2007117155A1 NZ 2007000044 W NZ2007000044 W NZ 2007000044W WO 2007117155 A1 WO2007117155 A1 WO 2007117155A1
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WO
WIPO (PCT)
Prior art keywords
zone
test
assay
complex
solid phase
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.)
Ceased
Application number
PCT/NZ2007/000044
Other languages
English (en)
Inventor
Keith Henderson
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
New Zealand Institute for Bioeconomy Science Ltd
Original Assignee
AgResearch Ltd
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 AgResearch Ltd filed Critical AgResearch Ltd
Publication of WO2007117155A1 publication Critical patent/WO2007117155A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • G01N33/54386—Analytical elements
    • G01N33/54387—Immunochromatographic test strips
    • G01N33/54388—Immunochromatographic test strips based on lateral flow
    • 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
    • G01N33/689—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 pregnancy or the gonads
    • 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
    • G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
    • G01N33/743—Steroid hormones
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/36—Gynecology or obstetrics
    • G01N2800/368—Pregnancy complicated by disease or abnormalities of pregnancy, e.g. preeclampsia, preterm labour

Definitions

  • the present invention relates to a solid-phase immuno-assay.
  • the present invention relates to a solid-phase immuno-assay and related methodology suitable for determining pregnancy in equities.
  • equines For ease of reference the present invention will now be described with specific reference to equines. However, it should be appreciated that this should not be seen as necessarily restricting the scope of the present invention.
  • the determination of pregnancy in equines is presently problematic. For example, determining pregnancy by internal methods such as palpation or ultrasound can often be a traumatic experience for a horse, and in some cases can cause physical injury, either of which may endanger the health of mare and foetus.
  • a solid phase immuno-assay in the form of: a test strip for determining the pregnancy status of equines, via an analyte present in urine, which includes: a solid support comprising: - a control zone; a test zone; and a sample zone; all of said zones being in fluid communication with each other; characterised in that: the control zone includes a ligand specific for a labelled-complex; the test zone includes the analyte or analogue thereof; the sample zone includes the labelled-complex specific for the analyte; and wherein the sample zone is located upstream of the test zone, which is upstream of the control zone.
  • the present invention will now be described in relation to equines. However this should not be seen as limiting the scope of the invention.
  • 'analyte' refers to oestrone sulphate (OS).
  • the term 'analogue' refers to a compound having a structure which is similar to oestrone sulphate.
  • the test-zone may use Oestrogen Bovine Serum Albumin conjugate as the analogue.
  • 'ligand' refers to a molecule which can bind specifically to another molecule.
  • the ligand may be an immuno-molecule. Most preferably, the ligand is an antibody.
  • the term 'immuno-molecule' refers to a molecule derived from an immune system which is capable of specifically binding to another molecule. It may include an antibody, antibody binding fragment, or a phage display molecule.
  • the term 'labelled-complex' refers to a complex of compounds which includes a tracer compound bound, or otherwise associated, with an immuno-molecule specific for oestrone sulphate, and may also include oestrone sulphate once bound to the immuno-molecule portion of the complex.
  • the labelled-complex may be a conjugated gold- oestrone sulphate monoclonal antibody complex (OS detection antibody complex).
  • the term 'antibody' refers to an immunoglobulin molecule having an amino acid sequence which specifically binds to an antigen, such as oestrone sulphate.
  • the term 'antibody' includes antibody binding fragments.
  • an antibody binding fragment may include, but not be limited to, an Fr, F(ab) 2 fragment, or ScFr molecule or the like.
  • the antibody may be a monoclonal antibody or a binding fragment thereof.
  • the term 'tracer compound' encompasses any compound which is capable of being visibly determined by the eye without the need for instrumentation.
  • solid support may come in a variety of different forms without departing from the scope of the present invention.
  • the solid support may be in the form of a cellulose ester.
  • the solid support may be a nitro-cellulose paper.
  • the solid support may have a surface area capable of supporting the ligand in a concentration of at least 0.5mg/ml and the analyte or an analogue thereof in a concentration of 0.5 to 10 mg/ml Most preferably the support may also have a surface area capable of supporting 0.02ml of the labelled- complex.
  • the surface area of the paper may be at least 25 mm x 5mm.
  • the test strip may be substantially housed within a body configured so that the control zone, test zone and sample zone are visible and/or accessible.
  • a sample of urine may be collected from a mare 100 days post-mating, and this is added to the sample-zone where it soaks into the solid-support and releases the conjugated gold-OS monoclonal antibody complex ("OS detection antibody complex”), which starts to migrate up the solid support.
  • OS detection antibody complex conjugated gold-OS monoclonal antibody complex
  • OS detection antibody When OS is present in the urine, this can become bound to the OS detection antibody. If OS is not present, or the OS concentration is low, the antigen-binding sites in the OS detection antibody do not become filled, and as a result when the OS detection antibody passes over the test-zone, some of it binds to the analyte analogue in the solid-support and a visible dot forms. This indicates the mare is not pregnant.
  • a solid phase immunoassay test may be prepared as follows:
  • Affinity purified IgG fraction of monoclonal antibody to oestrone sulphate is dissolved in 2mM sodium tetraborate (0.01 to 1 mg antibody/ml), and 0.5 ml added rapidly to 10 ml colloidal gold particles (40 nm) undergoing fast stirring.
  • the solution is left to stand for 1 hour at 26° C, then 1 ml of 1 % BSA in 2mM sodium tetraborate added and the incubation continued for a further 15 min.
  • the solution is centrifuged at 450O 9 (av) for 10 min at 4° C to pellet the conjugated gold-lgG particles.
  • PuraBind A-FP nitrocellulose membrane test platforms (75X300mm) are purchased from Whatman International Ltd. Solutions of oestrogen-bovine serum albumin conjugate (O-BSA;1 to 10 mg/ml) and anti-mouse IgG (1 mg/ml) are prepared in coating buffer (50 mmol/l carbonate-bicarbonate buffer, pH 9.6). A 'test' zone is generated by applying a line (1mm thick) of the O-BSA solution, 9mm from the bottom edge of the PuraBind A-FP membrane. Preparation of a Control Zone
  • a 'control' zone is generated by applying a line (1mm thick) of the anti-mouse IgG solution, 5mm above the test zone 'line'. After drying for 30 minutes at 37° C, the platform is immersed in blocking solution (aqueous 1% polyvinylpyrrolidone, average molecular weight 10 000, and 0.5% Triton X-100) for 30 minutes at room temperature. The platform is then washed with 5 changes of distilled water, dried at 37° C for 30 minutes and stored dry with silica gel desiccant at room temperature.
  • blocking solution aqueous 1% polyvinylpyrrolidone, average molecular weight 10 000, and 0.5% Triton X-100
  • the prepared PuraBind A-FP membrane platforms are cut into 75 X 5 mm strips, and the adhesive areas above and below the nitrocellulose membrane exposed by removing the silicone release tapes.
  • An absorbent sink (Whatman 17 ChR cellulose paper, 13x5 mm) is attached to the top of the membrane (2 mm overlap) and a prepared conjugate release pad to the bottom of the membrane (2 mm overlap).
  • a sample application pad (Whatman GF/D glass fibre paper, 17x5 mm) is attached fully overlapping the conjugate release pad.
  • Each assembled strip is placed inside a plastic housing cassette (5mm) and sealed inside an aluminium pouch together with a disposable exact volume plastic pipette (100 ⁇ l) and a silica gel pack (1g). The packaged devices are stored at 4 0 C.
  • the packaged device is allowed to warm to room temperature before being opened.
  • a 100 ⁇ l aliquot of mare test urine is added drop by drop into- the sample well of the plastic cassette using the exact volume pipette.
  • a line in the test zone and a line in the control zone appear in the cassette window between 5 and 10 minutes, with maximum colour intensity occurring within 20 minutes.
  • the colour intensity of the line in the test zone is inversely proportional to the concentration of oestrone sulphate in the urine sample.
  • the appearance of 2 visible lines is interpreted as indicative of non- pregnancy.
  • the appearance of a control line only is interpreted as indicative of pregnancy.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Immunology (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Hematology (AREA)
  • Urology & Nephrology (AREA)
  • Food Science & Technology (AREA)
  • General Physics & Mathematics (AREA)
  • Cell Biology (AREA)
  • Biotechnology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Pathology (AREA)
  • Endocrinology (AREA)
  • Gynecology & Obstetrics (AREA)
  • Pregnancy & Childbirth (AREA)
  • Reproductive Health (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

La présente invention concerne un essai immunologique en phase solide constitué d'une bandelette d'essai permettant de déterminer le statut gravidique d'équidés grâce à un analyte présent dans l'urine, ledit essai incluant : un support solide contenant une zone de régulation, une zone d'essai et une zone d'échantillon ; l'ensemble desdites zones se trouvant en communication fluidique mutuelle. L'essai est caractérisé en ce que la zone de régulation inclut un ligand spécifique d'un complexe marqué, la zone d'essai inclut l'analyte ou un analogue de celui-ci, et la zone d'échantillon inclut le complexe marqué, spécifique de l'analyte. La zone d'échantillon est positionnée en amont de la zone d'essai, laquelle se trouve en amont de la zone de régulation.
PCT/NZ2007/000044 2006-04-07 2007-03-07 Essai immunologique en phase solide Ceased WO2007117155A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
NZ54648506 2006-04-07
NZ546485 2006-04-07

Publications (1)

Publication Number Publication Date
WO2007117155A1 true WO2007117155A1 (fr) 2007-10-18

Family

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/NZ2007/000044 Ceased WO2007117155A1 (fr) 2006-04-07 2007-03-07 Essai immunologique en phase solide

Country Status (1)

Country Link
WO (1) WO2007117155A1 (fr)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020178857A1 (fr) * 2019-03-06 2020-09-10 Council Of Scientific & Industrial Research Dispositif non invasif à base de papier permettant la détection d'une grossesse chez le bétail et les buffles
RU2809759C2 (ru) * 2020-03-05 2023-12-18 Каунсел Оф Сайнтифик Энд Индастриал Рисерч Неинвазивное устройство на бумажной основе для определения беременности у крупного рогатого скота и буйволов

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6187598B1 (en) * 1987-04-27 2001-02-13 Conopco Inc. Capillary immunoassay and device therefor comprising mobilizable particulate labelled reagents

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6187598B1 (en) * 1987-04-27 2001-02-13 Conopco Inc. Capillary immunoassay and device therefor comprising mobilizable particulate labelled reagents

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
HENDERSON K. ET AL.: "A dipstick immunoassay to rapidly measure serum oestrone sulfate concentrations in horses", REPROD. FERTIL. DEV., vol. 12, no. 3-4, 2000, pages 183 - 189, XP009040014 *
MONFORT S.L. ET AL.: "Monitoring ovarian function and pregnancy by evaluating excretion of urinary oestrogen conjugates in semi-free-ranging Przewalski's horses (Equus przewalskii)", JOURNAL OF REPRODUCTION AND FERTILITY, vol. 91, no. 1, January 1991 (1991-01-01), pages 155 - 164 *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020178857A1 (fr) * 2019-03-06 2020-09-10 Council Of Scientific & Industrial Research Dispositif non invasif à base de papier permettant la détection d'une grossesse chez le bétail et les buffles
AU2020233121B2 (en) * 2019-03-06 2024-12-19 Council Of Scientific & Industrial Research A non-invasive paper based device for pregnancy detection in cattle and buffaloes
US12546787B2 (en) 2019-03-06 2026-02-10 Council Of Scientific & Industrial Research Non-invasive paper based device for pregnancy detection in cattle and buffaloes
RU2809759C2 (ru) * 2020-03-05 2023-12-18 Каунсел Оф Сайнтифик Энд Индастриал Рисерч Неинвазивное устройство на бумажной основе для определения беременности у крупного рогатого скота и буйволов

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