Microbe detection via hybridizing magnetic relaxation nanosensors

• US 9,109,249 B2
• Filed: 05/21/2012
• Issued: 08/18/2015
• Est. Priority Date: 05/19/2011
• Status: Expired due to Fees

• Current Assignee: University of Central Florida Research Foundation Inc. (State University System of Florida)
• Original Assignee: University of Central Florida Research Foundation Inc. (State University System of Florida)
• Inventors: Naser, Saleh, Perez, J. Manuel, Kaittanis, Charalambos
• Primary Examiner(s): Whisenant, Ethan C
• Application Number:
  US14/118,834
• Publication Number:
  US 20140220565A1
• Time in Patent Office: 1,184 Days
• Field of Search: 435/6.1, 435/283.1, 536/22.1, 536/23.1, 536/24.3
• US Class Current: 1/1
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/689 for bacteria
• CPC Class Codes: C12Q 2563/143 Magnetism, e.g. magnetic label
• CPC Class Codes: C12Q 2563/155 Particles of a defined size, e.g. nanoparticles
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode

Abstract

Disclosed herein are methods and materials for facilitating the detection of nucleic acid analytes of interest. Specifically exemplified herein are methods for detecting mycobacterial microorganisms, namely Mycobacterium avium spp. paratuberculosis. Also disclosed is new hybridizing magnetic relaxation nanosensor (hMRS) particularly adapted to detect a target nucleic acid analyte of interest.

Claims

1. A method of detecting a nucleic acid analyte of interest in a sample, the method comprising:

(a) contacting nucleic acid from a sample with a hybridizing magnetic relaxation nanosensor (hMRS) comprising a plurality of nucleic acid sequence probes configured to specifically hybridize to one unique sequence of the nucleic acid analyte of interest, so that only one hMRS hybridizes per nucleic acid analyte of interest; and

(b) detecting a change in the magnetic resonance signal from a hMRS hybridized to a nucleic acid analyte of interest in the nucleic acid from the sample;

thereby detecting the presence of the nucleic acid analyte of interest in the sample.

2. The method of claim 1, further comprising the step of isolating nucleic acid from the sample to obtain isolated nucleic acid.

3. The method of claim 2, wherein said contacting nucleic acid from the sample comprises contacting isolated nucleic acid with at least one hMRS.

4. The method of claim 1, wherein said sample is any sample harboring microorganisms from clinical or environmental sources.

5. The method of claim 1, wherein the nucleic acid analyte of interest is of a microorganism.

6. The method of claim 5, wherein the microorganism is a strain of bacteria.

7. The method of claim 6, wherein the bacteria is a Mycobacterium, and wherein the Mycobacterium is Mycobacterium avium spp. Paratuberculosis.

8. The method of claim 5, wherein said nucleic acid analyte of interest is DNA or RNA sequence of a microorganism.

9. The method of claim 8, wherein said nucleic acid analyte of interest is IS900 nucleotide sequence specific to MAP.

10. The method of claim 1, wherein said hMRS comprises a magnetic nanoparticle.

11. The method of claim 10, wherein said magnetic nanoparticle comprises a metal or metal oxide.

12. The method of claim 1, wherein said method can detect the presence of a nucleic acid analyte of interest within 90 minutes or less.

13. A hybridizing magnetic relaxation nanosensor (hMRS), comprising a magnetic nanoparticle associated with a plurality of nucleic acid probes configured to specifically hybridize to one unique sequence of a nucleic acid analyte of interest, so that only one hMRS hybridizes per nucleic acid analyte of interest.

14. The hMRS of claim 13, wherein said magnetic nanoparticle comprises a metal or metal oxide.

15. The hMRS of claim 13, wherein said magnetic nanoparticle comprises iron oxide.

16. The hMRS of claim 13, wherein said nucleic acid analyte of interest is of a microorganism.

17. The hMRS of claim 16, wherein said microorganism is a strain of bacteria.

18. The hMRS of claim 17, wherein said bacteria is a Mycobacterium, and wherein said Mycobacterium is Mycobacterium avium spp. Paratuberculosis.