EP0746768A1 - Procede de determination de la proteine c-reactive dans le lait pour le diagnostic de maladies inflammatoires des glandes mammaires et/ou controle de la qualite du lait et/ou des produits laitiers, et moyens pour la mise en oeuvre dudit procede - Google Patents

Procede de determination de la proteine c-reactive dans le lait pour le diagnostic de maladies inflammatoires des glandes mammaires et/ou controle de la qualite du lait et/ou des produits laitiers, et moyens pour la mise en oeuvre dudit procede

Info

Publication number
EP0746768A1
EP0746768A1 EP95910499A EP95910499A EP0746768A1 EP 0746768 A1 EP0746768 A1 EP 0746768A1 EP 95910499 A EP95910499 A EP 95910499A EP 95910499 A EP95910499 A EP 95910499A EP 0746768 A1 EP0746768 A1 EP 0746768A1
Authority
EP
European Patent Office
Prior art keywords
milk
determination
reactive protein
crp
mastitis
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.)
Withdrawn
Application number
EP95910499A
Other languages
German (de)
English (en)
Inventor
Wieland SCHRÖDL
Rudolf Kunze
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.)
Imtox Privatinstitut fur Immunbiologische Forschung GmbH
Original Assignee
Imtox Privatinstitut fur Immunbiologische Forschung GmbH
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 Imtox Privatinstitut fur Immunbiologische Forschung GmbH filed Critical Imtox Privatinstitut fur Immunbiologische Forschung GmbH
Publication of EP0746768A1 publication Critical patent/EP0746768A1/fr
Withdrawn legal-status Critical Current

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
    • G01N33/6893—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 diseases not provided for elsewhere
    • 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
    • G01N33/02—Food
    • G01N33/04—Dairy products
    • 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
    • G01N2333/4737—C-reactive protein

Definitions

  • the present invention relates to a method for the qualitative, semi-quantitative and quantitative determination of the C-reactive protein (CRP) from the milk of mammals for the diagnosis of inflammatory diseases of the mammary glands, in particular mastitis or testing the quality of milk or products obtained from milk , as well as means for implementation in practice.
  • CRP C-reactive protein
  • Mastitis is one of the most common diseases of the gonads.
  • Mastitis is an inflammatory disease of the milk-producing glands, e.g. B. due to a bacterial infection or mechanical irritation.
  • the disease causes a decrease in milk production and milk quality. Due to the deterioration in milk quality due to mastitis, for example, cattle milk is no longer for ; Suitable for calves.
  • mastitis for example, cattle milk is no longer for ; Suitable for calves.
  • a great deal of damage arises from the fact that milk from diseased animals, for example cattle, sheep or goats, may not be processed to produce foods, for example drinking milk or cheese, or may lead to defective products in the case of further processing.
  • the treatment of mastitis is mandatory and is usually done locally on the diseased organ. Difficulties are often the early diagnosis of mastitis and the observation of its course.
  • ⁇ antitrypsin and ⁇ 2 macrogiobulin. Both ⁇ -
  • All of these parameters are an expression of a local inflammatory reaction or disease of the mammary gland. The enumerated parameters have in common that they are only little for a routine laboratory medical companion examination to detect mastitis, e.g. B. that of cattle are suitable.
  • An indirect method for determining the number of lines is the detection of the cells by their DNA content, known as the Californian Mastitis Test (7), which, however, cannot discriminate between inflammation-relevant and non-inflammation-relevant cells.
  • the qualitative and quantitative detection of bacteria in the mastitis using microbiological tests is material-consuming and lengthy.
  • a reliable and fast method for diagnosing inflammatory diseases of the mammary glands, such as mastitis, using laboratory diagnostic methods is still not available in practice.
  • the CRP is a known acute phase protein. It is phylogenetically a very old protein and occurs in an identical or similar structure in practically all large groups of animals (3, 4). Its concentration can increase rapidly and very rapidly in inflammatory processes in the bloodstream (3, 4) and is particularly induced by bacterial infections. In inflammatory processes, CRP in the blood can rise relatively quickly. Under certain conditions, CRP is an important parameter for assessing the severity and course of inflammatory diseases (3, 4).
  • the detection of elevated CRP concentrations in mastitis milk samples is surprising, since CRP can only be detected in milk after appropriate preparation. As a result of the preparation of the samples, the CRP becomes accessible and can be recognized in the detection system.
  • the preparation takes place without clarification of the milk and precipitation of milk components. This preparation is carried out, for example, by adding a preparation solution to which a detergent, for example Tween 20 and a chelating agent for polyvalent cations, for example ethyldi ⁇ itrilotetraacetic acid (EDTA), is preferably added.
  • a detergent for example Tween 20
  • a chelating agent for polyvalent cations for example ethyldi ⁇ itrilotetraacetic acid (EDTA)
  • EDTA ethyldi ⁇ itrilotetraacetic acid
  • the principle of the method is that specific CRP-erken ⁇ en ⁇ es antiserum or antibody / mixture bound to a solid phase, after binding of the analyte (CRP), a second antiserum or antibody / mixture, which is preferably conjugated to an enzyme, at the still free binding sites of the analyte bin ⁇ et.
  • CRP analyte
  • a second antiserum or antibody / mixture which is preferably conjugated to an enzyme
  • Figure 3 shows the comparison of CRP in milk from healthy and mastitis-sick cows; the Mann-Whitney test was significant at p ⁇ 0.001.
  • C-reactive protein or a fragment of the C-reactive protein in the milk of mammals the milk being from mammals suffering from an inflammatory disease of the mammary gland, preferably mastitis.
  • C-reactive protein or a fragment of the C-reactive protein in the milk of mammals or in milk-containing products and in combination with other chemical, biochemical or immunological laboratory parameters Determination of C-reactive protein or a fragment of the C-reactive protein in the milk of mammals as an accompanying laboratory test for therapies of mastitis or other mammary gland diseases.
  • C-reactive protein or a fragment of the C-reactive protein in the milk of mammals whereby detection methods for the determination of CRP are used using polyclonal antisera or monoclonal or oligoclonal antibodies.
  • Determination of C-reactive protein or a fragment of the C-reactive protein in the milk of mammals the detection being carried out by means of an immunoassay, preferably an enzyme immunoassay (EIA).
  • an immunoassay preferably an enzyme immunoassay (EIA).
  • milk or milk products can be prepared using chelating agents and detergents or mixtures composed thereof.
  • a 96-well microtiter plate which has previously been coated with an antiseru against CRP and can be stored after drying until the time of the CRP determination, is prepared for the determination by washing once, preferably with 0.1 M phosphate-buffered saline. This is done by adding 100 ⁇ l, if possible with a multichannel micro pipette, to each well and after 1 minute the contents are removed by knocking out.
  • the preparation to be examined for example the milk, is mixed with the preparation solution, which consists of 5 mM EDTA and Tween 20 0.05% (vol. / Vol.) In phosphate-buffered saline (50 ⁇ l sample + 450 ⁇ l preparation solution) and mixed thoroughly. 100 ⁇ l of the sample to be examined are filled into 1 well of the microtiter plate. The plate loaded with the samples is sealed with an adhesive film to prevent evaporation of the sample and incubated for 60 minutes at room temperature. After the incubation, the samples are knocked out and the plate is washed by filling it three times with 250 ⁇ l of a 0.1 M phosphate-buffered saline solution per well and then knocking out the plate.
  • the preparation solution which consists of 5 mM EDTA and Tween 20 0.05% (vol. / Vol.)
  • phosphate-buffered saline 50 ⁇ l sample + 450 ⁇ l preparation solution
  • a second anti-CRP antiserum which is conjugated with peroxidase, is now added (100 ⁇ l per well).
  • the plate is sealed with an adhesive film and incubated for 60 minutes at room temperature. After the incubation period, the plate is knocked out again and washed three times as described above.
  • 100 ⁇ l of substrate solution are added to each well, which contains 3,3 ' -5,5 ' tetramethyibenzidine (TMB) and H 2 O 2 dissolved in a phosphate citrate buffer. After 1 5 minutes, 50 ⁇ l of a stop solution (2 MH 2 SO 4 ) is added to each well to terminate the enzyme-substrate reaction.
  • TMB 3,3 ' -5,5 ' tetramethyibenzidine
  • the amount of the converted substrate correlates with the amount of the detected CRP and enables the CRP content of the sample to be estimated optically.
  • photometric measurement of the optical density at 450 nm and taking into account a standard CRP curve quantitative determination of the CRP in the sample is possible.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Food Science & Technology (AREA)
  • Immunology (AREA)
  • General Physics & Mathematics (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Medicinal Chemistry (AREA)
  • Pathology (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Analytical Chemistry (AREA)
  • Hematology (AREA)
  • Cell Biology (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

L'invention concerne un procédé de diagnostic et/ou de surveillance du traitement d'une maladie inflammatoire de(s) glande(s) mammaire(s) chez un mammifère, de préférence une mastite, qui comprend l'étape consistant à déterminer la protéine C-réactive ou un fragment de cette dernière dans le lait dudit mammifère.
EP95910499A 1994-02-21 1995-02-17 Procede de determination de la proteine c-reactive dans le lait pour le diagnostic de maladies inflammatoires des glandes mammaires et/ou controle de la qualite du lait et/ou des produits laitiers, et moyens pour la mise en oeuvre dudit procede Withdrawn EP0746768A1 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DE4405893 1994-02-21
DE19944405893 DE4405893A1 (de) 1994-02-21 1994-02-21 Verfahren zur Bestimmung von C-reaktiven Protein in der Milch zur Diagnostizierung von entzündlichen Erkrankungen der Milchdrüse und/oder Prüfung der Qualität von Milch und/oder Milchprodukten und Mittel zu seiner Durchführung
PCT/EP1995/000587 WO1995022767A1 (fr) 1994-02-21 1995-02-17 Procede de determination de la proteine c-reactive dans le lait pour le diagnostic de maladies inflammatoires des glandes mammaires et/ou controle de la qualite du lait et/ou des produits laitiers, et moyens pour la mise en oeuvre dudit procede

Publications (1)

Publication Number Publication Date
EP0746768A1 true EP0746768A1 (fr) 1996-12-11

Family

ID=6511040

Family Applications (1)

Application Number Title Priority Date Filing Date
EP95910499A Withdrawn EP0746768A1 (fr) 1994-02-21 1995-02-17 Procede de determination de la proteine c-reactive dans le lait pour le diagnostic de maladies inflammatoires des glandes mammaires et/ou controle de la qualite du lait et/ou des produits laitiers, et moyens pour la mise en oeuvre dudit procede

Country Status (3)

Country Link
EP (1) EP0746768A1 (fr)
DE (1) DE4405893A1 (fr)
WO (1) WO1995022767A1 (fr)

Families Citing this family (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB9924180D0 (en) 1999-10-12 1999-12-15 Univ Glasgow Assays for mastitis
DE10106253C2 (de) * 2001-02-10 2003-09-18 Oliver Lehnsdorf Verfahren zur Qualitätsuntersuchung von Milch

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO9522767A1 *

Also Published As

Publication number Publication date
DE4405893A1 (de) 1995-09-14
WO1995022767A1 (fr) 1995-08-24

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