# US 9,518,296 B2 · Methods, compositions, and kits for detecting protein aggregates > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 9,518,296 B2 - Document ID: US9518296B2 - Status: Granted - Kind: B2 - Application number: 13/975,056 - Filed: 2013-08-23 - Published: 2014-05-15 - Granted: 2016-12-13 - Earliest claimed: 2006-05-03 - Assignees: Applied Biosystems, LLC - Applicants: Applied Biosystems LLC - Inventors: David Ruff; Mark Shannon; Kenneth Livak; Karl Guegler; Kevin Hennessy - Web page: https://patentgrep.com/patent/US9518296B2 ## CPC - C12Q 1/6876: Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes - C07K 14/47: from mammals - C07K 14/4711: {Alzheimer's disease; Amyloid plaque core protein} - C12Q 1/6816: characterised by the detection means (C12Q1/6804 takes precedence) - C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) ## Abstract The present teachings provide methods, compositions, and kits for detecting the presence of protein aggregates. In some embodiments, the protein aggregate is treated with a labeled precursor, and the labeled precursor is incorporated into the protein aggregate to form a labeled protein aggregate. The labeled protein aggregate is then measured, thus detecting the presence of the protein aggregate. In some embodiments, the labeled protein aggregate is detected by interaction of labeled precursors, for example by a proximity ligation assay. ## Claims ### Claim 1 A kit for detecting a protein aggregate in a sample comprising: a first soluble precursor molecule; a second soluble precursor molecule; and an oligonucleotide splint, wherein the first soluble precursor molecule is attached to a first oligonucleotide, and wherein the second soluble precursor molecule is attached to a second oligonucleotide, and wherein the first and second soluble precursor molecules are protein moieties of the protein aggregate prior to aggregation. ### Claim 2 (depends on 1) The kit according to claim 1 further comprising a ligase. ### Claim 3 (depends on 1) The kit according to claim 1 further comprising reagents for a polymerase chain reaction (PCR), wherein the reagents for the PCR comprise a first primer and a second primer, wherein the first primer corresponds to the first oligonucleotide of the first soluble precursor molecule and wherein the second primer corresponds to the second oligonucleotide of the second soluble precursor molecule. ### Claim 4 (depends on 1) The kit of claim 1, where the first soluble precursor molecule and the second soluble precursor molecule are Sup35. ### Claim 5 (depends on 1) The kit of claim 1, where the first soluble precursor molecule and the second soluble precursor molecule are beta amyloid protein. ### Claim 6 (depends on 1) The kit of claim 1, where the first soluble precursor molecule and the second soluble precursor molecule are huntingtin. ### Claim 7 (depends on 1) The kit of claim 1, where the first soluble precursor molecule and the second soluble precursor molecule are alpha-synuclein. ### Claim 8 (depends on 1) The kit of claim 1, where the first soluble precursor molecule and the second soluble precursor molecule are bovine prion protein. ### Claim 9 (depends on 1) The kit of claim 1, where the reagents for PCR further comprise one or more of deoxynucleotide phosphates, thermostable DNA polymerase, one or more PCR buffers. ### Claim 10 (depends on 1) The kit of claim 1, wherein the first and second soluble precursor molecules are correctly folded proteins that are not yet in the conformation present in a protein aggregate. ### Claim 11 (depends on 1) The kit of claim 1, wherein the protein aggregate is a naturally occurring homo-multimeric protein complex. ### Claim 12 (depends on 1) The kit of claim 1, wherein the protein aggregate comprises abnormal mis-folded proteins. ### Claim 13 (depends on 1) The kit of claim 1, wherein the protein aggregate comprises aggregated prion particles. ### Claim 14 (depends on 13) The kit of claim 13, wherein the aggregated prion particles comprise yeast prion proteins selected from Sup35, Ure2 and Rnq1, human prion proteins, bovine prion proteins, or sheep prion proteins. ### Claim 15 (depends on 1) The kit of claim 1, wherein the protein aggregate detected by the kit is selected from aggregated yeast prion proteins, human prion proteins, beta amyloid protein, tau proteins, huntingtin protein, alpha-synuclein protein, or bovine prion protein.