Method and kit for extracting prion protein
AssigneeThe United States of America as represented by the Secretary of Agriculture
InventorsMary Jo Schmerr, Andrew J. Alpert
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Abstract
A method for extracting prion protein from a biological material, e.g., an animal tissue or product. In a specific example, abnormal prion protein is extracted from homogenized sheep brain with hexafluoro-2-propanol. The hexafluoro-2-propanol is separated from the aqueous brain preparation by increasing the ionic strength of the aqueous solution. Prion protein in the organic extract can be further purified, or the extract can be tested, e.g., by immunoassay, for the presence of prion protein, and more particularly abnormal prion protein. The extraction process permits testing for the presence of abnormal prior protein, e.g., for diagnosis of transmissible spongiform encephalopathies (TSE).
Claims26
A method for extracting abnormal prion protein from a biological material suspected of containing abnormal prion protein, said method comprising:
- (a) incubating a mixture of an extraction solvent and an isotonic or hypotonic aqueous preparation of biological material under conditions effective to extract abnormal prion protein from said biological material into said extraction solvent, wherein said extraction solvent is;
- (i) a polar organic solvent in which said abnormal prion protein is soluble, and
- (ii) miscible with a non-lyotropic aqueous solution but immiscible with a lyotropic aqueous solution;
- and
- (b) increasing lyotropic activity of the mixture to separate said extraction solvent from said aqueous preparation of said biological material to yield extraction solvent containing any abnormal prion protein from said biological material.
A method for extracting abnormal prion protein from a biological material suspected of containing abnormal prion protein, said method comprising:
- (a) incubating a mixture of about equal amounts of hexafluoro-2-propanol and a non-lyotropic aqueous preparation of biological material under conditions effective to extract abnormal prion protein from said biological material into said hexafluoro-2-propanol;
- (b) adding about an equal volume of a solution of 0.5M sodium sulfate to said mixture to separate said hexafluoro-2-propanol from said aqueous preparation of said biological material to yield extraction solvent containing any abnormal prion protein from said biological material.
A kit for isolating abnormal prion protein from a biological sample, said kit comprising:
- (a) an extraction solvent, wherein the extraction solvent is
- (i) a polar organic solvent in which abnormal prion protein is soluble, and
- (ii) miscible with a hypotonic or isotonic aqueous solution but immiscible with a lyotropic aqueous solution;
- and
- (b) a lyotropic salt or aqueous lyotropic salt solution to add to an aqueous preparation of a biological sample so that said organic solvent becomes immiscible with said aqueous preparation.
A kit for detecting the presence of abnormal prion protein from a biological sample, said kit comprising
- (a) an extraction solvent which is
- (i) a polar organic solvent in which abnormal prion protein is soluble, and
- (ii) miscible with a non-lyotropic aqueous solution but immiscible with a lyotropic aqueous solution;
- (b) a lyotropic salt or aqueous lyotropic salt solution to add to an aqueous preparation of a biological sample so that said extraction solvent becomes immiscible with said aqueous preparation;
- and
- (c) an abnormal prion protein detection assay.
Dependencies are inferred from the claim text.