Method and apparatus for identifying analyte-containing samples using single-read determination of analyte and process control signals

• US 9,822,397 B2
• Filed: 06/30/2011
• Issued: 11/21/2017
• Est. Priority Date: 06/30/2010
• Status: Active Grant

• Current Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Original Assignee: Gen-Probe Incorporated (Hologic Incorporated)
• Inventors: Joo, Sunghae A., Dockter, Janel M.
• Primary Examiner(s): Dauner, Joseph G
• Application Number:
  US13/173,558
• Publication Number:
  US 20120003646A1
• Time in Patent Office: 2,336 Days
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 2537/165 Mathematical modelling, e.g. logarithm, ratio
• CPC Class Codes: C12Q 2545/101 with an internal standard/control
• CPC Class Codes: C12Q 2545/114 involving a quantitation step

Abstract

Apparatus and method for detecting analyte in a reaction mixture comprising an internal control. The invention is illustrated using amplification and detection of nucleic acids, where the amplification reaction comprises an exogenous internal control. Magnitude of the detection signal serves as the variable for identifying invalid trials, identifying valid trials that are negative for analyte, and identifying trials that are positive for analyte. Detection of a signal indicating probe hybridization can be used for assigning the presence or absence of analyte nucleic acid in validated reactions using only a single detectable label, and without distinguishing the proportion of signal contributed by internal control probe binding, and analyte probe binding.

Claims

1. A method for determining the presence or absence of an analyte nucleic acid in a sample, the method comprising the steps of:

(a) forming a reaction mixture comprising a sample suspected of containing an analyte nucleic acid, an internal control (IC) nucleic acid that is distinct from the analyte nucleic acid, and reagents sufficient to synthesize amplicons using the analyte nucleic acid and the IC nucleic acid as templates;

(b) after step (a), subjecting the reaction mixture to amplification reaction conditions, the amplification reaction conditions permitting analyte amplicons to be synthesized from the analyte nucleic acid, if present in the sample, and IC amplicons to be synthesized from the IC nucleic acid;

(c) exposing the reaction mixture to an analyte probe and an IC probe under conditions permitting the analyte probe to bind to the analyte amplicons, if any, and the IC probe to bind to the IC amplicons synthesized in the reaction mixture during step (b), wherein the analyte probe and the IC probe include first and second detectable labels, respectively, wherein signals generated by the first and second detectable labels are indistinguishable from each other, and wherein the analyte probe is specific for the analyte amplicons and the IC probe is specific for the IC amplicons; and

(d) detecting a combined signal generated by the first and second detectable labels, the combined signal being indicative of the presence or absence of the analyte nucleic acid in the sample, wherein the analyte probe and the IC probe are both free in solution during the detecting step, and wherein the detecting step is performed at a constant temperature.

2. The method of claim 1, wherein the first and second detectable labels are identical to each other.

3. The method of claim 1, wherein each of the first and second detectable labels is a chemiluminescent label.

4. The method of claim 1, wherein each of the first and second detectable labels is a fluorescent label.

5. The method of claim 1, wherein each of the analyte probe and the IC probe is a self-hybridizing probe having an interacting label pair when the self-hybridizing probe is in a close conformation.

6. The method of claim 5, wherein the self-hybridizing probe is a molecular torch or a molecular beacon.

7. The method of claim 1, wherein the detecting step is performed using a single detection channel in a detection device.