Electrophoretics Limited

University of Geneva

The United States of America as Represented by the Department of Health

Bio-Rad Laboratories, Inc.

Universitè De Genève

Bayer Intellectual Property Gmbh

Electrophorectics Limited

Bayer Schering Pharma Ag

Denis Francois Hochstrasser

Jean-Charles Sanchez

Gerhard Koenig

Oezkan Yalkinoglu

Willy Vincent Bienvenut

Laure Allard

Elisabeth Guillaume

Catherine Gabrielle Zimmerman

Pierre Lescuyer

Odile Carrette

Abraham Patchornik

Carl R Merril

Özkan Yalkinoglu

Catherine Gabrielle Zimmermann

Carl R Merrill

Gerhard König

Jean Charles Sanchez

Özkan A Yalkinoglu

A brain damage-related disorder is diagnosed in a subject by detecting at least one polypeptide, or a variant or mutant thereof, in a sample of body fluid taken from the subject, wherein the polypeptide is one for which the level is either increased or decreased in cerebrospinal fluid from deceased patients compared to cerebrospinal fluid from healthy donors.

Diagnostic method for brain damage-related disorders

A method for assessing the state of Alzheimer's disease in patients is disclosed. A method for monitoring the progression of Alzheimer's disease in patients is also disclosed. The method applies detection of specific peptide markers, e.g., using mass spectrometric analysis.

Biomarkers for diagnosing alzheimer's disease

A brain damage-related disorder is diagnosed in a subject by detecting at least one polypeptide, or a variant or mutant thereof, selected from A-FABP, E-FABP, PGP 9.5, GFAP, Prostaglandin D synthase, Neuromodulin, Neurofilament L, Calcyphosine, RNA binding regulatory subunit, Ubiquitin fusion degradation protein 1 homolog, Nucleoside diphosphate kinase A, Glutathione S tranferase P, Cathepsin D, DJ-1 protein, Peroxiredoxin 5 and Peptidyl-prolyl cis-trans isomerase A (Cyclophilin A) in a sample of body fluid taken from the subject.

Stroke is diagnosed in a subject by determining the concentration of at least one polypeptide selected from Apo C-III, Serum Amyloid A, Apo C-I, Antithrombin III fragment and Apo A-I in a sample of body fluid taken from the subject.

Diagnostic method for stroke

Heart and brain fatty acid binding proteins (H-FABP, B-FABP) are markers for stroke. The invention provides a diagnostic assay for either of these markers, preferably by ELISA using a anti-H-FABP or B-FABP antibody. Since H-FABP is also a marker for acute myocardial infarction (AMI), to distinguish stroke from AMI requires an assay specific to AMI, e.g. using troponin-1 or CK-MB as a marker, also to be carried out.

Diagnostic assay for stroke

Heart and brain fatty acid binding proteins (H-FABP, B-FABP) are markers for TSEs, especially CJD. The invention provides a diagnostic assay for either of these markers, preferably by enzyme immunoassay using a specific antibody thereto. Since H-FABP is also a marker for acute myocardial infarction (AMI), to distinguish CJD from AMI requires an assay specific to AMI, e.g. using troponin-1 or CK-MB as a marker, also to be carried out.

Diagnostic assay for transmissible spongiform encephalopathies

Heart and brain fatty acid binding proteins (H-FABP, B-FABP) are markers for stroke. The invention provides a diagnostic assay for either of these markers, preferably by ELISA using anti-H-FABP or B-FABP antibody. Since H-FABP is also a marker for acute myocardial infarction (AMI), to distinguish stroke from AMI requires an assay specific to AMI, e.g. using troponin-I or creatine kinase-MB as a marker, also to be carried out.

Polypeptides which have been separated by gel electrophoresis can be identified or characterized by a procedure which has two main stages. In the first stage the gel is digested with a polypeptide-cleaving agent such as an enzyme. This produces mainly large fragments which, in the second stage are electroblotted through a hydrophilic membrane on which is immobilized another polypeptide-cleaving reagent such as an enzyme onto a hydrophobic member, typically a membrane, e.g. of PVDF. The resulting fragments, usually peptides, are identified, preferably by MALDI-TOF MS, or a property may be determined, e.g. by interaction with an antibody.

Method and kit for identifying or characterizing polypeptides

Method of identifying polypeptides

Polypeptides separated on a gel are identified by cleaving the separated polypeptides with an immobilized cleaving reagent such as an enzyme, and transferring the fragments to a hydrophobic collection layer where they are analyzed. A hydrophilic membrane containing an immobilized protease such as trypsin is provided between an electrophoresis gel and a hydrophobic membrane to form an electroblotting &ldquo;sandwich&rdquo;. Polypeptides separated on the gel by electrophoresis are electroblotted from the gel through the hydrophilic membrane where they are cleaved by the protease into fragments, and the fragments are collected on the hydrophobic membrane where they are identified such as by MALDI-TOF MS analysis. From identification of the fragments, the polypeptide from which they came is identified. The hydrophilic membrane may be provided with functional groups to which the protease is immobilized by covalent bonding. Free residual functional groups not bound to the protease are capped to prevent their reaction with the polypeptide. The functional groups may be activated carbonyl groups, carboxylic acid groups or carboxylic acid derivative groups capable of reacting with an amino group of an enzyme. A kit for use in carrying out the electroblotting is formed containing the cleaving reagent immobilized on the hydrophilic membrane, and the hydrophobic collection layer. The immobilized cleaving reagent and hydrophobic layer may be in separate containers or in the same container.

Kit for electroblotting polypeptides separated on an electrophoresis gel

A polyacrylamide gel comprising acrylamide and diacrylylpiperazine. A method for providing a polyacrylamide gel comprising employing a catalyst system which comprises dimethylpiperazine, sodium thiosulfate or a mixture thereof and ammonium or potassium persulfate. The use of the gel as the matrix in a silver staining procedure for the detection of biomaterials, provides for the obtainment of reduced background staining.

Catalyst for preparing polyacrylamide gel which improves the detection

Amine-acryloyl and -methacryloyl derivatives and 1,6-heptadiene-4-ol can be used to provide crosslinked polyacrylamide gels which are useful in protein and nucleic acid separation and detection. The gels crosslinked with the compounds of the present invention exhibit much less background silver staining than previously known acrylamide gels.

Polyacrylamide gels with improved detection of protein

Polyacrylamide gels containing chaotropic agents such as urea as solubilizing agents for protein or nucleic acid separations are rendered stable for prolonged periods of time by the use of certain diacrylyl compounds with tertiary amide groups as cross linking agents. These compounds have the general structure ##STR1## where the R groups are generally saturated hydrocarbon groups and R.sup.4 through R.sup.9 may also be hydrogen. When used as cross linkers, these compounds have the further advantage of preventing urea crystallization at lowered temperatures.

Stable polyacrylamide gels containing chaotropic agents

A strip gel and a slab gel are combined in a single gel enclosure or on a single gel backing for two-dimensional electrophoresis. The enclosure is comprised of a single pair of support plates, and a removable electrically insulating region is included to separate the gels. The region is occupied either by air or an electrically nonconductive material which can be removed without disturbing either gel. In use, a sample is loaded onto one end of the strip gel and an electric potential is imposed across the strip gel for the first dimension separation while the strip gel is electrically insulated from the slab gel. The insulating region is then removed and the two gels placed in electrical contact for the second dimension separation.

Growing community of inventors

Geneva, Switzerland

Denis Francois Hochstrasser