Sensitive and rapid determination of antimicrobial susceptibility

• US 8,460,887 B2
• Filed: 02/08/2010
• Issued: 06/11/2013
• Est. Priority Date: 07/12/2003
• Status: Active Grant

• Current Assignee: Accelerate Diagnostics, Inc.
• Original Assignee: Accelerate Diagnostics, Inc.
• Inventors: Goldberg, David A., Howson, David C., Metzger, Steven W., Buttry, Daniel A., Saavedra, Steven Scott
• Primary Examiner(s): LAM, ANN Y
• Application Number:
  US12/702,210
• Publication Number:
  US 20100136570A1
• Time in Patent Office: 1,219 Days
• Field of Search: None
• US Class Current: 435/7.2
• CPC Class Codes: B82Y 10/00 Nanotechnology for information processing, storage or transmission, e.g. quantum computing or single electron logic
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
• CPC Class Codes: B82Y 20/00 Nanooptics, e.g. quantum optics or photonic crystals
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
• CPC Class Codes: C12Q 1/18 Testing for antimicrobial activity of a material
• CPC Class Codes: G01N 33/5438 Electrodes
• CPC Class Codes: G01N 33/561 Immunoelectrophoresis
• CPC Class Codes: G01N 33/569 for microorganisms, e.g. protozoa, bacteria, viruses

Abstract

The present invention relates to moving microorganisms to a surface, where they are grown in the presence and absence of antimicrobials, and by monitoring the growth of the microorganisms over time in the two conditions, their susceptibility to the antimicrobials can be determined. The microorganisms can be moved to the surface through electrophoresis, centrifugation or filtration. When the movement involves electrophoresis, the presence of oxidizing and reducing reagents lowers the voltage at which electrophoretic force can be generated and allows a broader range of means by which the target can be detected. Monitoring can comprise optical detection, and most conveniently includes the detection of individual microorganisms. The microorganisms can be stained in order to give information about their response to antimicrobials.

Claims

1. A method of detecting the response of a test microorganism to a condition, comprising:

capturing the microorganism on a surface of a substrate using an applied force, wherein the surface has an affinity for the microorganism;

detecting at a first time a property of the microorganism on the surface with a camera;

placing the microorganism in a condition;

detecting at a second time the property of the microorganism on the surface with the camera; and

determining the response of the microorganism to the condition by the amount of difference of the property of the microorganism between the first time and the second time.

2. The method of claim 1, wherein the microorganism is selected from the set consisting of bacterium, mycoplasma, protozoan, fungus, and plant cell and animal cell.

3. The method of claim 1, wherein the affinity of the surface is conferred by a cationic polymer.

4. The method of claim 1, wherein the applied force comprises electrophoresis.

5. The method of claim 4, wherein the electrolyte in which electrophoresis occurs comprises an oxidizing agent and a reducing agent, wherein the potential at which the electrophoresis occurs is below 2 volts.

6. The method of claim 1, wherein the applied force comprises filtration.

7. The method of claim 1, wherein the applied force comprises magnetic force, and a paramagnetic tag is bound to the microorganism in order to confer responsiveness of the microorganism to a magnetic field.

8. The method of claim 1, wherein the force comprises centrifugation.

9. The method of claim 1, wherein the force comprises dielectrophoresis.

10. The method of claim 1, wherein the property comprises the amount of stain that is detected.

11. The method of claim 10, wherein the stain is selected from the set consisting of vital stain and mortal stain.

12. The method of claim 1, wherein the property comprises the size of the organism.

13. The method of claim 1, wherein the property comprises the presence of a second microorganism at the second time adjacent to the location of the test microorganism, wherein the second microorganism is not present at the first time.

14. The method of claim 1, wherein the condition comprises the presence of an agent selected from the set consisting of antibiotic, bacteriophage, animal virus, double-stranded RNA, DNA, anticancer drug, antifungal drug, antimalarial drug, and antiprotozoal drug.

15. The method of claim 1, wherein determining at the first time additionally comprises locating the microorganism on the surface with the camera and storing the location of the microorganism in a storage medium, whereby the microorganism is located at the second time by retrieving the location from the storage medium.