Diagnostic and sample preparation devices and methods

• US 9,580,742 B2
• Filed: 03/12/2012
• Issued: 02/28/2017
• Est. Priority Date: 03/10/2011
• Status: Active Grant

• Current Assignee: General Atomics, Inc.
• Original Assignee: Xagenic Inc
• Inventors: Kelley, Shana O., Bortolin, Susan, Orton, Reginald James McKenzie, Wiechula, Stefan Christopher
• Primary Examiner(s): CROW, ROBERT THOMAS
• Application Number:
  US14/003,709
• Publication Number:
  US 20140170646A1
• Time in Patent Office: 1,814 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: C12N 1/066 by physical methods
• CPC Class Codes: C12N 13/00 Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
• CPC Class Codes: C12N 15/1003 Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2523/307 Denaturation or renaturation by electric current/voltage
• CPC Class Codes: C12Q 2525/204 specific length of the oligonucleotides
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer
• CPC Class Codes: C12Q 2565/607 being a sensor, e.g. electrode
• CPC Class Codes: C12Q 2565/629 being a microfluidic device

Abstract

Contemplated methods and devices are drawn to preparation and analysis of analytes from biological samples. In a preferred embodiment the analytes are nucleic acids that are both released from biological compartment present in the sample and fragmented through the use of a voltage potential applied to a pair of electrodes. The nucleic acids thus prepared are subsequently characterized.

Claims

1. A method of analyzing a biological sample, the method comprising:

transferring the biological sample to an extraction zone, wherein the extraction zone comprises a pair of electrodes;

selecting a first electrical signal to apply to the pair of electrodes; and

applying the first electrical signal to the pair of electrodes, such that a plurality of nucleic acids are released from the biological sample and fragmented within the extraction zone to produce a plurality of fragmented nucleic acids,

wherein selecting the first electrical signal comprises selecting, based on a desired average length in bases of the plurality of fragmented nucleic acids, at least one of a magnitude of a voltage applied to the pair of electrodes, a duration of a voltage pulse applied to the pair of electrodes and a frequency at which a voltage pulse is applied to the pair of electrodes.

2. The method of claim 1, wherein the average length in bases of the fragmented nucleic acids is equal to or less than 50% of the average length in bases of the plurality of nucleic acids prior to fragmentation.

3. The method of claim 1, wherein the plurality of fragmented nucleic acids have an average length of equal to or less than 200 bases.

4. The method of claim 1, wherein the first electrical signal is selected such that the average length of the plurality of fragmented nucleic acids is less than or equal to a predetermined average length.

5. The method of claim 4, wherein selecting the first electrical signal further comprises selecting the overall treatment time.

6. The method of claim 4, wherein selecting the first electrical signal is effective to produce the plurality of fragmented nucleic acids to be capable of hybridization to a solid-phase bound capture probe in a hybridization time that is at least 25% less than a corresponding hybridization time of the plurality of nucleic acids prior to fragmentation.

7. The method of claim 1, further comprising contacting the biological sample in the extraction zone with an enhancing reagent.

8. The method of claim 7, wherein the enhancing reagent is selected from the group consisting of a metal ion, a free radical promoting compound, a chaotropic salt, an ionic detergent, and a nonionic detergent.

9. The method of claim 1, further comprising moving the biological sample containing the fragmented nucleic acids to an analysis zone and applying a second electrical signal to a sensing electrode and a reference electrode.

10. The method of claim 9, further comprising measuring a charge or current flow between the sensing electrode and the reference electrode to quantify or ascertain the presence of a target nucleic acid in the biological sample.

11. The method of claim 10, wherein the sensing electrode is a nanostructured microelectrode, and wherein the nanostructured microelectrode is spiky, rough or fractal.