IP Library › Granted Patent US 12,385,080
Granted Patent B2
US 12,385,080 · App. 16/575,149 · Granted Aug 12, 2025

Detection of analytes and nucleic acids

Inventors: Mark Shannon (San Francisco, CA); David Ruff (Oxford, GB)
Assignee: Applied Biosystems, LLC
C12Q1/6804
View Patent ↗
Loading inventors, assignments & file history…
Monitor This Case
Get email alerts when status or documents change.
Order Certified Copies
Most orders are placed with the USPTO same day — all within 24 business hours.
Order via The Patent Place →
Pre-filled with this patent's details
Quick Facts
Patent No.
US 12,385,080
App. No.
16/575,149
Granted
Aug 12, 2025
Kind
B2
Abstract

Methods of detecting at least one analyte and at least one nucleic acid in a sample are provided. Reagents for carrying out the methods are also provided.

Claims (36)

1. A method of detecting at least one target analyte and at least one target nucleic acid in a cell, comprising:

lysing the cell in a multifunctional lysis buffer to produce a cell lysate;

incubating the cell lysate with:

i) at least one proximity detection probe set including at least two proximity detection probes, wherein each of the at least two proximity detection probes comprises at least one analyte binding moiety and at least one oligonucleotide moiety;

ii) at least one splint oligonucleotide; and

iii) at least one ligase;

such that at least one ligated proximity detection probe set is formed;

incubating the cell lysate with at least one protease;

detecting the at least one target analyte by detecting the at least one ligated proximity detection probe set; and

detecting the at least one target nucleic acid,

wherein the detecting the at least one ligated proximity detection probe set includes removing an aliquot of the cell lysate from a vessel, prior to incubating the cell lysate with at least one protease, and

the detecting the at least one target nucleic acid is performed using a remaining portion of the cell lysate in the vessel, after incubating the cell lysate with at least one protease.

2. The method of claim 1 , wherein at least one target analyte is a protein.

3. The method of claim 1 , wherein the multifunctional lysis buffer is capable of lysing the cell without substantially degrading the at least one target nucleic acid.

4. The method of claim 3 , wherein less than 50% of the at least one target nucleic acid is degraded.

5. The method of claim 1 , wherein the multifunctional lysis buffer is capable of lysing the cell while maintaining a structure of the at least one target analyte.

6. The method of claim 1 , wherein at least 50% of the at least one target analyte maintains the structure.

7. The method of claim 1 , wherein the cell lysate is incubated with the at least one proximity detection probe set, the at least one splint oligonucleotide and the at least one ligase at a temperature between 0° C. and 45° C.

8. The method of claim 1 , wherein the cell lysate is incubated with the at least one protease at a temperature of at least 55° C.

9. The method of claim 8 , further comprising wherein the cell lysate is further incubated with the at least one protease at a temperature of at least 65° C.

10. The method of claim 1 , wherein the detecting the at least one target analyte includes performing at least one of reverse transcription and real-time polymerase chain reaction.

11. A method of detecting at least one target analyte and at least one target nucleic acid in a cell, comprising:

lysing the cell in a multifunctional lysis buffer to produce a cell lysate;

incubating the cell lysate with at least one proximity detection probe set such that at least one hybridized proximity detection probe set is formed, wherein the at least one proximity detection probe set comprises at least two proximity detection probes, and wherein each of the at least two proximity detection probes comprises at least one analyte binding moiety and at least one oligonucleotide moiety;

incubating the cell lysate with at least one protease;

detecting the at least one target analyte by detecting the at least one hybridized proximity detection probe set; and

detecting the at least one target nucleic acid,

wherein the detecting the at least one ligated proximity detection probe set includes removing an aliquot of the cell lysate from a vessel, prior to incubating the cell lysate with at least one protease, and

the detecting the at least one target nucleic acid is performed using a remaining portion of the cell lysate in the vessel, after incubating the cell lysate with at least one protease.

12. The method of claim 11 , wherein at least one target analyte is a protein.

13. The method of claim 11 , wherein the incubating the cell lysate with at least one proximity detection probe set comprises incubating the cell lysate with at least one splint oligonucleotide and the at least one proximity detection probe set.

14. The method of claim 11 , wherein the detecting the at least one hybridized proximity detection probe set occurs prior to the incubating of the cell lysate with the at least one protease.

15. The method of claim 14 , wherein the detecting the at least one hybridized proximity detection probe set includes using an aliquot of the cell lysate, after the incubating of the cell lysate with the at least one hybridized proximity detection probe set.

16. The method of claim 11 , further comprising:

incubating the cell lysate with an enzyme suitable for ligating ends of the at least one oligonucleotide moiety, after the incubating of the cell lysate with the at least one proximity detection probe set.

17. The method of claim 11 , wherein the detecting the at least one target analyte includes performing at least one of reverse transcription and real-time polymerase chain reaction.

Assignments (4)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 16, 2020
From: SHANNON, MARK E.; RUFF, DAVID W.
To: APPLERA CORPORATION
Reel/Frame 051540/0778 →
CHANGE OF NAME Recorded Jan 16, 2020
From: APPLERA CORPORATION
To: APPLIED BIOSYSTEMS, INC.
Reel/Frame 051540/0840 →
MERGER Recorded Jan 16, 2020
From: APPLIED BIOSYSTEMS, INC.
To: ATOM ACQUISITION CORPORATION WITH/AND INTO APPLIED BIOSYSTEMS, INC.
Reel/Frame 051540/0897 →
MERGER AND CHANGE OF NAME Recorded Jan 16, 2020
From: ATOM ACQUISITION CORPORATION WITH/AND INTO APPLIED BIOSYSTEMS, INC.; APPLIED BIOSYSTEMS, INC.
To: APPLIED BIOSYSTEMS, LLC
Reel/Frame 051540/0939 →
Continuity (5)
Continuation 14744643 · Jun 19, 2015
Continuation 13194735 · Jul 29, 2011
Continuation 11888656 · Jul 31, 2007
Provisional Application 60835118 · Aug 1, 2006
Related Publication 20200102592A1 · Apr 2, 2020
References Cited (114)
US 4988617A · Landegren et al. · 1991 [cited by applicant]
US 5362853A · Kuga et al. · 1994 [cited by applicant]
US 5534410A · Tjian et al. · 1996 [cited by applicant]
US 5665539A · Sano et al. · 1997 [cited by applicant]
US 5674987A · Gray · 1997 [cited by applicant]
US 5863726A · Harley et al. · 1999 [cited by applicant]
US 5874260A · Cleuziat et al. · 1999 [cited by applicant]
US 5876924A · Zhang et al. · 1999 [cited by applicant]
US 5879885A · Becker · 1999 [cited by applicant]
US 5935825A · Nishimura et al. · 1999 [cited by applicant]
US 6037130A · Tyagi et al. · 2000 [cited by applicant]
US 6124133A · Taylor et al. · 2000 [cited by applicant]
US 6210915B1 · Shay et al. · 2001 [cited by applicant]
US 6333157B1 · Miller-Jones et al. · 2001 [cited by applicant]
US 6342389B1 · Cubicciotti · 2002 [cited by applicant]
US 6511809B2 · Baez et al. · 2003 [cited by applicant]
US 6762027B2 · Greenfield et al. · 2004 [cited by applicant]
US 7863050B2 · Domanico et al. · 2011 [cited by applicant]
US 8012685B2 · Shannon et al. · 2011 [cited by applicant]
US 20010018183A1 · Bao et al. · 2001 [cited by applicant]
US 20010018513A1 · Baker · 2001 [cited by applicant]
US 20020001823A1 · Southan et al. · 2002 [cited by applicant]
US 20020064779A1 · Landegren et al. · 2002 [cited by applicant]
US 20020102591A1 · Sorge · 2002 [cited by applicant]
US 20020177139A1 · Greenfield et al. · 2002 [cited by applicant]
US 20020187475A1 · Coticone et al. · 2002 [cited by applicant]
US 20030027234A1 · Pandian et al. · 2003 [cited by applicant]
US 20030091992A1 · Lorincz et al. · 2003 [cited by applicant]
US 20030119152A1 · Bleicher et al. · 2003 [cited by applicant]
US 20030148525A1 · Wang · 2003 [cited by applicant]
US 20040197769A1 · Wong et al. · 2004 [cited by applicant]
US 20040248103A1 · Feaver et al. · 2004 [cited by applicant]
US 20050003361A1 · Fredriksson · 2005 [cited by applicant]
US 20050006239A1 · Amshey et al. · 2005 [cited by applicant]
US 20050009036A1 · Montesclaros et al. · 2005 [cited by applicant]
US 20050009045A1 · Greenfield et al. · 2005 [cited by applicant]
US 20050019893A1 · Huletsky et al. · 2005 [cited by applicant]
US 20050181508A1 · Fredriksson et al. · 2005 [cited by applicant]
US 20050282158A1 · Landegren · 2005 [cited by applicant]
US 20060063763A1 · Schall et al. · 2006 [cited by applicant]
US 20060078882A1 · Zetter et al. · 2006 [cited by applicant]
US 20060141556A1 · Jeong et al. · 2006 [cited by applicant]
US 20060194216A1 · Willey et al. · 2006 [cited by applicant]
US 20060214104A1 · Pope et al. · 2006 [cited by applicant]
US 20060223071A1 · Wisniewski et al. · 2006 [cited by applicant]
US 20060241072A1 · Baker · 2006 [cited by applicant]
US 20070026430A1 · Andersen et al. · 2007 [cited by applicant]
US 20070041960A1 · Freeman et al. · 2007 [cited by applicant]
US 20070111231A1 · Magness et al. · 2007 [cited by applicant]
US 20070134648A1 · Soskic · 2007 [cited by applicant]
US 20070281367A1 · Hennessy et al. · 2007 [cited by applicant]
US 20080050731A1 · Agnew et al. · 2008 [cited by applicant]
US 20080064071A1 · Hogrefe et al. · 2008 [cited by applicant]
US 20080187924A1 · Korfhage et al. · 2008 [cited by applicant]
US 20080220979A1 · Wang et al. · 2008 [cited by applicant]
US 20090246859A1 · Domanico et al. · 2009 [cited by applicant]
US 20100055678A1 · Jaatinen et al. · 2010 [cited by applicant]
US 20100311041A1 · Brillowska-Dabrowska · 2010 [cited by applicant]
US 20110244457A1 · Nadeau et al. · 2011 [cited by applicant]
US 20120135885A1 · Lam et al. · 2012 [cited by applicant]
US 20150132743A1 · Egidio et al. · 2015 [cited by applicant]
EP 1249500A1 · 2002 [cited by applicant]
EP 1566437A1 · 2005 [cited by applicant]
WO WO9932654A1 · 1999 [cited by applicant]
WO WO0056920A1 · 2000 [cited by applicant]
WO WO0161037A1 · 2001 [cited by examiner]
WO WO03044231A1 · 2003 [cited by applicant]
WO WO2003064605A2 · 2003 [cited by applicant]
WO WO03104385A1 · 2003 [cited by applicant]
WO WO2004027082A2 · 2004 [cited by applicant]
WO WO2004094456 · 2004 [cited by examiner]
WO WO2004094456A2 · 2004 [cited by examiner]
WO WO2005040194A2 · 2005 [cited by applicant]
WO WO2005087917A2 · 2005 [cited by applicant]
WO WO2005123963A2 · 2005 [cited by applicant]
WO WO2008016644A1 · 2008 [cited by applicant]
Ambion, “Quantitate Stem Cell Pluripotency Proteins Using TaqMan Real-Time PCR Assays”, TechNotes Newsletter, vol. 16, No. 3, 2011, pp. 24-25. [cited by applicant]
Amendment and Response filed Aug. 13, 2007 in U.S. Appl. No. 11/428, 191. [cited by applicant]
Bjarnegard, M. et al., Endothelium-specific ablation of PDGFB leads to pericyte loss and glomerular, cardiac and placental abnormalities, Development, 131 (8), 2004, pp. 1847-1857. [cited by applicant]
Cao, W., “Recent Developments in Ligase-mediated Amplification and Detection”, Trends in Biotechnoloav. vol. 22, No. 1, Elsevier Ltd., 2004, pp. 38-44. [cited by applicant]
Chen et al., “Discordant Protein and mRNA Expression in Lung Adenocarcinomas”, Molecular & Cellular Proteomics, vol. 1.4, 2002, pp. 304-313. [cited by applicant]
Di Giusto, D. et al., “Construction, Stability, and Activity of Multivalent Circular Anticoagulant Aptamers”, Journal of Biological Chemistry. 2004, vol. 279(45): pp. 46483-46489. [cited by applicant]
Di Giusto, D. et al., “Proximity Extension of Circular DNA Aptamers with Real-Time Protein Detection”, Nucleic Acids Research, vol. 33, No. 6: e64, 2005, 7 pages. [cited by applicant]
Eis, P. et al., “An Invasive Cleavage Assay for Direct Quantitation of Specific RNAs.”, Nature Biotechnology. 2001, vol. 19: pp. 673-676. [cited by applicant]
EP 07810985.7 EP Examination Report, mailed Jan. 5, 2010. [cited by applicant]
European Search Report for Application No. 17187196.5, mailed Mar. 6, 2018, 8 pages. [cited by applicant]
Extended European Search Report for Application No. 12150643.0, mailed Jan. 16, 2013, 14 pages. [cited by applicant]
Fredriksson, et al., “Protein detection using proximity-dependent DNA ligation assays”, Nature Biotechnology. vol. 20, May 1, 2002, pp. 473-477. [cited by applicant]
Griffin et al., Detection of hemizygosity in Hermansky-Pudlak syndrome by quantitative real-timePCR. Clinical Genetics 68 : 23 (2005). [cited by applicant]
Gullberg, et al., “A Sense of Closeness; Protein Detection by Proximity Ligation”, Current Opinion in Biotechnology, vol. 14, No. 1, 2003, pp. 82-86. [cited by applicant]
Gullberg, et al., “Cytokine Detection by Antibody-based Proximity Ligation”, Proceedings of National Academy of Science, vol. 101, No. 22, Jun. 1, 2004, pp. 8420-8424. [cited by applicant]
Gustafsdottir, S. et al., “Detection of Individual Microbial Pathogens by Proximity Ligation”, Clinical Chemistry. 2006, vol. 52(6): pp. 1152-1160. [cited by applicant]
Gustafsdottir, S. et al., “Proximity ligation assays for sensitive and specific protein analyses”, Analytical Biochemistry, vol. 345, Elsevier Inc., 2005, pp. 2-9. [cited by applicant]
Han, M. et al., “Quantum-dot-tagged Microbeads for Multiplexed Optical Coding of Biomolecules,”, Nature Biotechnology, vol. 19, Jul. 2001, 631-635. [cited by applicant]
International Search Report and Written Opinion dated Dec. 20, 2007, in PCT Application No. PCT/US2007/017187. [cited by applicant]
Johansson, M.K . . . , Choosing Reporter-Quencher Pairs for Efficient Quenching Through Formation of Intramolecular Dimers. Methods in Molecular Biology vol. 335, Ed. by Didenko (2006). [cited by applicant]
Lee et al.,Seven-Color, Homogeneous Detection of Six PCR Products. Biotechniques 27(2) : 342 (1999). [cited by applicant]
Neophytou, P. et al., “Development of a Procedure for the Direct Cloning of T-Cell Epitopes Using Bacterial Expression Systems”, Journal of Immunological Methods. vol. 196, Issue 1, 1996, pp. 63-72. [cited by applicant]
Newton, C. et al., “In der Reihe FOCUS sind auBerdem erschienen”, PCR, Speiktrum Akademischer Verlag, Heidelberg, DE, 1994, 7 pages. [cited by applicant]
Office Action mailed Apr. 12, 2007, in U.S. Appl. No. 11/428,191. [cited by applicant]
Office Action mailed Oct. 22, 2007, in U.S. Appl. No. 11/428,191. [cited by applicant]
Oliver, C.N. et al., “A proteolytic artifact associated with lysis of bacteria by egg white lysozyme”, Proc. Natl. Acad. Sci. USA vol. 80, 1983, pp. 2156-2160. [cited by applicant]
Pai, et al., “Proximity Litigation Assays with Peptide Conjugate ‘Burrs’ for the Sensitive Detection of Spores”, Nucleic Acids Research, vol. 33, No. 18, 2005, e162, pp. 1-7. [cited by applicant]
Partial European Search Report for Application No. 12150643.0, mailed Sep. 26, 2012, 8 pages. [cited by applicant]
PCT/US2007/017187 “International Preliminary Report on Patentability,” issued Feb. 3, 2009. [cited by applicant]
Sambrook, Fritsch, Maniatis: “Molecular Cloning: A Laboratory Manual, Second Edition”, Jan. 1, 1989 (Jan. 1, 1989), Cold Spring Harbor Laboratory Press, XP002682632. [cited by applicant]
Sambrook, J. et al., “Isolation of Cytoplasmic RNA from Mammalian Cells”, Molecular Cloning: A Laboratory Manual, 2nd Edition, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, 1989, pp. 7.12-7.15. [cited by applicant]
Sano, T. et al., “Immuno-PCR: Very Sensitive Antigen Detection by Means of Specific Antibody-DNA Conjugates”, Science, 1992, vol. 258, pp. 120-122. [cited by applicant]
Sims, Paul et al., “Immunopolymerase Chain Reaction Using Real-Time Polymerase Chain Reaction for Detection”, Analytical Biochemistry, 281, 2000, 230-232. [cited by applicant]
Spiro et al.,A Bead-Based Method for Multiplexed Identification and Quantitation of DNA Sequences Using Flow Cytometry. Applied and Environmental Microbiology 66(10) : 4258 (2000). [cited by applicant]
Stratagene, Catalog (1988), “Stragagene Cloning Systems: Tools and Technology for Life Sciences”, p. 39. Published by Stratgene, 11011 North Torrey Pines Road, La Jolla, CA 92037, USA. [cited by applicant]
Table reprinted from Calbiochem-Novabiochem Manuscript/Paper entitled Detergents :A guide to the properties and uses in biological systems (2001). [cited by applicant]
Vuillard, L. et al., “Non-detergent Sulfobetaines: a new class of mild solubilization agents for protein purification”, Biochemical Journal, 305, 1995, pp. 337-343. [cited by applicant]
Ward et al., Detection of Vibrio parahaemolyticus in Shellfish by Use of Multiplexed Real-Time PCR with TaqMan Fluorescent Probes. Applied and Enviromental Microbiology 72(3) : 2031 (2006). [cited by applicant]