EP0122478A2
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EP
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A method for the preparation of monoclonal antibody with specificity for crosslinked fibrin derivatives and an assay procedure using said antibody
Assignee: Agen Biomedical Ltd
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Filing: 1984-03-16
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Granted: —
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Expires: 2004-03-16
Abstract
Abstract The invention relates to a method of preparation of monoclonal antibody derived from a crosslinked fibrin derivative including the steps of: (i) obtaining a crosslinked fibrin derivative or extract containing same; and (ii) forming antibody to said derivative or extract by cloning antibody producing cells from an animal having been administered thereto said derivative or extract. There is also provided a screening assay for use in the abovementioned method which includes the steps of: (A) coating a surface with antigen selected from (i) crosslinked fibrin derivative; (ii) extract containing said derivative; or (iii) fibrinogen degradation product. (B) contacting said antigen with monoclonal antibody derived from crosslinked fibrin derivative; and (C) subjecting the complex formed in step (A) to a signal amplification step. There is also disclosed an assay procedure for detection of the presence of a crosslinked fibrin derivative in an animal body fluid including the steps of: (i) contacting monoclonal antibody prepared from a crosslinked fibrin derivative with a fluid sample suspected of containing an antigen derived from a crosslinked fibrin derivative or comprising a crosslinked fibrin derivative perse; and (ii) subjecting the complex formed in step (1) to a signal amplification step.
Inventors
- 👤 Peter Gregory Bundesen
- 👤 Dennis Brian Rylatt
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-02 05:38
CPC Classifications
C; C12; C12P; C12P21/00; G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/86; A; A61; A61K; A61K39/00; A61K39/395; C07; C07K; C07K16/00; C07K16/18; G01N2333/00; G01N2333/435; G01N2333/745; G01N2333/75
Claims Analysis
Total Claims: 20
Claim 1
Independent Claim
52 words
A method of preparation of monoclonal antibody derived from a crosslinked fibrin derivative including the steps of: (i) obtaining a crosslinked fibrin derivative or extract containing same; and (ii) forming antibody to said derivative or extract by cloning antibody producing cells from an animal having been administered thereto said derivative or extract.
Claim 2
Independent Claim
52 words
A method of preparation of monoclonal antibody derived from a crosslinked fibrin derivative including the steps of: (i) obtaining a crosslinked fibrin derivative or extract containing same; and (ii) forming antibody to said derivative or extract by cloning antibody producing cells from an animal having been administered thereto said derivative or extract.
Claim 3
Dependent Claim
38 words
A method as claimed in claim 1 wherein said extract is obtained from plasmic degradation of fibrin clots or by simultaneous action of thrombin, Factor XIII and plasmin or fibrinogen with transient clot formation and subsequent clot lysis.
Claim 4
Dependent Claim
30 words
A method as claimed in claim 1 wherein said extract is prepared by subjecting animal body fluid containing said crosslinked fibrin derivative to an electrophoresis, chromatographic or other fractionation technique.
Claim 5
Independent Claim
65 words
A method as claimed in any preceding claim wherein in step (ii) antigenic extract containing said crcsslinked fibrin derivative is administered to a mouse which is subsequently killed and the spleen subsequently removed for further purification to isolate lymphocytic or spleen white blood cells which are subsequently fused to mouse myeloma cells to form a hybridoma cell which may be cloned or recloned as desired.
Claim 6
Dependent Claim
29 words
A method as claimed in claim 4 wherein said hybridoma cells are cultivated in well plates whereby hybridoma cell suspensions are placed in each well with cell cultivation media.
Claim 7
Dependent Claim
63 words
A screening assay for use in the method of claim 1 including the steps of: (A) coating a surface with antigen selected from (i) crosslinked fibrin derivative; (ii) extract containing said derivative; or (iii) fibrinogen degradation product. (B) contacting said antigen with monoclonal antibody derived from crosslinked fibrin derivative; and (C) subjecting the complex formed in step (A) to a signal amplification step.
Claim 8
Dependent Claim
29 words
An assay as claimed in claim 6 wherein in step (A) a well plate is utilized in which antigen is added to each well prior to said monoclonal antibody.
Claim 9
Dependent Claim
71 words
8 . An assay procedure for detection of the presence of a crosslinked fibrin derivative in an animal body fluid including the steps of: (i) contacting monoclonal antibody prepared from a crosslinked fibrin derivative with a fluid sample suspected of containing an antigen derived from a crosslinked fibrin derivative or comprising a crosslinked fibrin derivative per se; and (ii) subjecting the complex formed in step (i) to a signal amplification step.
Claim 10
Dependent Claim
71 words
8 . An assay procedure for detection of the presence of a crosslinked fibrin derivative in an animal body fluid including the steps of: (i) contacting monoclonal antibody prepared from a crosslinked fibrin derivative with a fluid sample suspected of containing an antigen derived from a crosslinked fibrin derivative or comprising a crosslinked fibrin derivative per se; and (ii) subjecting the complex formed in step (i) to a signal amplification step.
Claim 11
Dependent Claim
18 words
9 . An assay procedure as claimed in claim 8 wherein the crosslinked fibrin derivative is D dimer.
Claim 12
Dependent Claim
42 words
An assay procedure as claimed in claim 8 or 9 wherein a first monoclonal antibody in step (i) is bound or captured to an antigen in said body fluid which is subsequently tagged in step (ii) by a labelled second monoclonal antibody.
Claim 13
Dependent Claim
23 words
An assay procedure as claimed in claim 10 wherein said first monoclonal antibody is monospecific and said second monoclonal antibody is pan specific.
Claim 14
Dependent Claim
23 words
An assay procedure as claimed in claim 1 0 wherein said first monoclonal antibody is monospecific and said second monoclonal antibody is monospecific.
Claim 15
Dependent Claim
23 words
An assay procedure as claimed in claim 10 wherein said first monoclonal antibody is pan specific and said second monoclonal antibody is monospecific.
Claim 16
Dependent Claim
33 words
An assay procedure as claimed in any one of claims 10 to 13 wherein the second monoclonal antibody is labelled with an enzyme which is subsequently reacted with a substrate for said enzyme.
Claim 17
Dependent Claim
24 words
An assay procedure as claimed in any one of claims 10 to 13 wherein the second monoclonal antibody is labelled with a radioactive species.
Claim 18
Dependent Claim
37 words
An assay procedure in claim 8 or 9 wherein the monoclonal antibody in step (i) is initially coupled to latex beads before being contacted with said fluid sample and subsequently being checked for agglutination in step (ii).
Claim 19
Independent Claim
162 words
A method of detection of crosslinked fibrin derivative in a body fluid including the steps of: (i) immunizing an animal with a crosslinked fibrin derivative or extract containing same; (ii) removing a spleen from the animal; (iii) treating the spleen to form a cell suspension; (iv) purifying the cell suspension to isolate spleen white blood cells or lymphocytes. (v) forming hybridoma cells containing as one component said spleen white blood cells or lymphocytes; (vi) cloning or recloning said hybridoma cells using appropriate cell feeder layers; (vii) carrying out screening assays with antigen selected from crosslinked fibrin derivative or extract containing same or fibrinogen degredation product so as to isolate hybridoma cells which produce monoclonal antibody specific to a crosslinked fibrin derivative; (viii) contacting a fluid sample suspected of containing crosslinked fibrin derivative or antigen derived therefrom with monoclonal antibody prepared from hybridoma cells isolated after step (vii), and (ix) subjecting the complex formed in step (viii) to a signal amplification step .
Claim 20
Independent Claim
162 words
A method of detection of crosslinked fibrin derivative in a body fluid including the steps of: (i) immunizing an animal with a crosslinked fibrin derivative or extract containing same; (ii) removing a spleen from the animal; (iii) treating the spleen to form a cell suspension; (iv) purifying the cell suspension to isolate spleen white blood cells or lymphocytes. (v) forming hybridoma cells containing as one component said spleen white blood cells or lymphocytes; (vi) cloning or recloning said hybridoma cells using appropriate cell feeder layers; (vii) carrying out screening assays with antigen selected from crosslinked fibrin derivative or extract containing same or fibrinogen degredation product so as to isolate hybridoma cells which produce monoclonal antibody specific to a crosslinked fibrin derivative; (viii) contacting a fluid sample suspected of containing crosslinked fibrin derivative or antigen derived therefrom with monoclonal antibody prepared from hybridoma cells isolated after step (vii), and (ix) subjecting the complex formed in step (viii) to a signal amplification step .
References Cited (Backward Citations)
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