NMR SYSTEMS AND METHODS FOR THE RAPID DETECTION OF ANALYTES

• US 20120100546A1
• Filed: 10/22/2010
• Published: 04/26/2012
• Est. Priority Date: 10/22/2010
• Status: Active Grant

• Current Assignee: T2 Biosystems, Inc
• Original Assignee: T2 Biosystems, Inc
• Inventors: Audeh, Mark John, Kumar, Sonia, Wellman, Parris, Lowery, Thomas Jay Jr., Blanco, Matthew, Chepin, James Franklin, Demas, Vasiliki, Dhanda, Rahul, Fritzemeier, Marilyn Lee, Koh, Isaac, Neely, Lori Anne, Mozeleski, Brian, Plourde, Daniella Lynn, Rittershaus, Charles William
• Granted Patent: US 8,563,298 B2
• Time in Patent Office: Days
• US Class Current: 435/6.12
• CPC Class Codes: B82Y 25/00 Nanomagnetism, e.g. magnetoimpedance, anisotropic magnetoresistance, giant magnetoresistance or tunneling magnetoresistance
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/6827 for detection of mutation or polymorphism
• CPC Class Codes: C12Q 1/689 for bacteria
• CPC Class Codes: C12Q 1/6895 for plants, fungi or algae
• CPC Class Codes: C12Q 1/701 Specific hybridization probes
• CPC Class Codes: C12Q 2563/143 Magnetism, e.g. magnetic label
• CPC Class Codes: C12Q 2565/633 NMR
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: G01N 24/08 by using nuclear magnetic resonance G01N24/12 takes precedence
• CPC Class Codes: G01N 27/745 for detecting magnetic beads used in biochemical assays concerning the assays G01N33/54326; sensors therefor G01R33/1269; automatic analysers therefor G01N35/0098
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/5434 using magnetic particle immunoreagent carriers which constitute new materials per se
• CPC Class Codes: G01R 33/093 using multilayer structures, e.g. giant magnetoresistance sensors thin magnetic films H01F10/00
• CPC Class Codes: G01R 33/1269 of molecules labeled with magnetic beads magnetic particles for bio assay G01N33/54326
• CPC Class Codes: G01R 33/281 Means for the use of in vitro contrast agents G01R33/282 takes precedence; involving use of a contrast agent in MR imaging G01R33/5601; in vivo contrast agents A61K49/0002
• CPC Class Codes: G01R 33/30 Sample handling arrangements, e.g. sample cells, spinning mechanisms
• CPC Class Codes: G01R 33/302 Miniaturized sample handling arrangements for sampling small quantities, e.g. flow-through microfluidic NMR chips
• CPC Class Codes: G01R 33/448 Relaxometry, i.e. quantification of relaxation times or spin density G01R33/50 takes precedence
• CPC Class Codes: Y10T 436/24 Nuclear magnetic resonance, electron spin resonance or other spin effects or mass spectrometry

Abstract

This invention features systems and methods for the detection of analytes, and their use in the treatment and diagnosis of disease.

Claims

1-98. -98. (canceled)

99. A composition comprising:

(a) a liquid sample containing an analyte; and

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 950 nm, a T₂ relaxivity per particle of from 1×

10⁴ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and binding moieties specific to the analyte are conjugated to their surface,

wherein the analyte from the sample is bound to at least one of the binding moieties on at least one magnetic particle.

100. A composition comprising:

(a) a liquid sample suspected of containing a Candida nucleic acid; and

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 700 nm to 1200 nm, a T₂ relaxivity per particle of from 1×

10⁹ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and having a first probe and a second probe conjugated to their surface selected so that the first probe is operative to bind to a first segment of the Candida nucleic acid and the second probe is operative to bind to a second segment of the Candida nucleic acid.

103. The composition of any one of claims 100-102, wherein the liquid sample is derived from a whole blood sample.

101. A composition comprising:

(a) a liquid sample suspected of containing creatinine;

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 699 nm, a T₂ relaxivity per particle of from 1×

10⁸ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and creatinine antibodies conjugated to their surface; and

(c) a multivalent binding agent bearing a plurality of creatinine conjugates designed to form aggregates with the population of magnetic particles in the absence of creatinine.

104. The composition of claim 101 or 102, wherein the multivalent binding agent comprises a polymeric scaffold.

102. A composition comprising:

(a) a liquid sample suspected of containing tacrolimus;

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 699 nm, a T₂ relaxivity per particle of from 1×

10⁸ to 1×

10¹² and creatinine antibodies conjugated to their surface; and

(c) a multivalent binding agent bearing a plurality of tacrolimus conjugates designed to form aggregates with the population of magnetic particles in the absence of tacrolimus.

105. A composition, comprising:

a) a whole blood sample, or a component thereof;

b) a forward primer comprising the oligonucleotide sequence 5′

-GGC ATG CCT GTT TGA GCG TC-3′

;

c) a reverse primer comprising the oligonucleotide sequence 5′

-GCT TAT TGA TAT GCT TAA GTT CAG CGG GT-3′

;

d) a thermal stable polymerase; and

e) deoxynucleotide triphosphates, buffer, and magnesium.

106. A composition, comprising:

(a) a liquid sample containing one or more nucleic acid amplicons generated from an amplification reaction; and

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 1200 nm, a T₂ relaxivity per particle of from 1×

10⁴ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and having a first probe and a second probe conjugated to their surface selected so that the first probe is operative to bind to a first segment of the nucleic acid amplicons and the second probe is operative to bind to a second segment of the nucleic acid amplicons.

107. A composition, comprising:

(a) a liquid sample containing Candida nucleic acid amplicons generated from an amplification reaction; and

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 1200 nm, a T₂ relaxivity per particle of from 1×

10⁴ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup> mM<sup>−

1</sup>s<sup>−

1</sup>, and having a first probe and a second probe conjugated to their surface selected so that the first probe is operative to bind to a first segment of the Candida nucleic acid amplicons and the second probe is operative to bind to a second segment of the Candida nucleic acid amplicons.

108. The composition of claim 107, wherein said magnetic particles have a mean diameter of from 700 nm to 1200 nm and a T₂ relaxivity per particle of from 1×

10⁹ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>.

109. The composition of claim 107, the binding of the first probe to a first segment of the Candida amplicon and the binding of the second probe to a second segment of the Candida amplicon form magnetic particle-amplicon aggregates.

110. The composition of claim 107, wherein the Candida nucleic acid amplicons are amplified from Candida albicans, and wherein the first probe comprises the oligonucleotide sequence:

111. The composition of claim 107, wherein the Candida nucleic acid amplicons are amplified from Candida krusei, and wherein the first probe comprises the oligonucleotide sequence:

112. The composition of claim 107, wherein the Candida nucleic acid amplicons are amplified from Candida glabrata, and wherein the first probe comprises the oligonucleotide sequence:

113. The composition of claim 107, wherein the Candida nucleic acid amplicons are amplified from Candida parapsilosis or Candida tropicalis, and wherein the first probe comprises the oligonucleotide sequence:

114. A composition comprising:

(a) a liquid sample containing nucleic acid amplicons generated from an amplification reaction;

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 1200 nm, a T₂ relaxivity per particle of from 1×

10⁴ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and having a first probe and a second probe conjugated to their surface and selected to hybridize to a first segment and a second segment of the nucleic acid amplicons to form aggregates, the aggregates comprising the magnetic particles and the nucleic acid amplicons.

115. The composition of claim 114, wherein said magnetic particles have a mean diameter of from 700 nm to 1200 nm and a T₂ relaxivity per particle of from 1×

10⁹ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>.

116. A composition comprising:

(a) a liquid sample containing Candida nucleic acid amplicons generated from an amplification reaction;

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 1200 nm, a T₂ relaxivity per particle of from 1×

10⁴ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and having a first probe and a second probe conjugated to their surface and selected to hybridize to a first segment and a second segment of the Candida nucleic acid amplicons to form aggregates, the aggregates comprising the magnetic particles and the Candida nucleic acid amplicons.

117. The composition of claim 116, wherein said magnetic particles have a mean diameter of from 700 nm to 1200 nm and a T₂ relaxivity per particle of from 1×

10⁹ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>.

118. The aggregate of claim 116, wherein the Candida nucleic acid amplicons are amplified from Candida albicans, and wherein the first probe comprises the oligonucleotide sequence:

119. The aggregate of claim 116, wherein the Candida nucleic acid amplicons are amplified from Candida krusei, and wherein the first probe comprises the oligonucleotide sequence:

120. The aggregate of claim 116, wherein the Candida nucleic acid amplicons are amplified from Candida glabrata, and wherein the first probe comprises the oligonucleotide sequence:

121. The aggregate of claim 116, wherein the Candida nucleic acid amplicons are amplified from Candida parapsilosis or Candida tropicalis, and wherein the first probe comprises the oligonucleotide sequence:

122. A composition comprising:

(a) a liquid sample optionally comprising an analyte;

(b) within the liquid sample from 1×

10⁶ to 1×

10¹³ magnetic particles per milliliter of the liquid sample, the magnetic particles having a mean diameter of from 150 nm to 699 nm, a T₂ relaxivity per particle of from 1×

10⁸ to 1×

10¹² mM<sup>−

1</sup>s<sup>−

1</sup>, and binding moieties specific to the analyte are conjugated to their surface; and

(c) within the liquid sample a multivalent binding agent bearing a plurality of analyte conjugates capable of forming aggregates with the population of magnetic particles in the absence of the analyte.

123. A method of detecting the presence of an analyte, comprising:

(a) combining a liquid sample containing an analyte with a population of magnetic particles having a mean diameter of from 150 nm to 1200 nm and binding moieties specific to a first binding site on the analyte are conjugated to their surface;

(b) exposing the sample to a magnetic field to capture or concentrate the magnetic particles and the analyte;

(c) exposing the concentrated magnetic particles to a surface derivitized with ligands complementary to a second binding site on the analyte thereby forming magnetic particle-analyte-ligand complexes;

(d) removing unbound magnetic particles to produce a population of analyte-bound magnetic particles;

(e) placing the analyte-bound magnetic particles in a well having an RF coil disposed about the well, the RF coil configured to detect a signal produced by exposing the liquid sample to a bias magnetic field created using one or more magnets and an RF pulse sequence;

(f) exposing the sample to a bias magnetic field and an RF pulse sequence;

(g) following step (f), measuring the T2 relaxation signal; and

(h) on the basis of the result of step (g), determining the concentration of analyte in the liquid sample.