Rapid cell purification systems

• US 9,714,420 B2
• Filed: 01/21/2016
• Issued: 07/25/2017
• Est. Priority Date: 03/07/2011
• Status: Active Grant

• Current Assignee: Accelerate Diagnostics, Inc.
• Original Assignee: Accelerate Diagnostics, Inc.
• Inventors: Hance, Kenneth Robert, Metzger, Steven W.
• Primary Examiner(s): Andres, Janet L
• Assistant Examiner(s): SNYDER, STUART
• Application Number:
  US15/003,604
• Publication Number:
  US 20160138067A1
• Time in Patent Office: 551 Days
• Field of Search: None
• CPC Class Codes: B03C 2201/18 Magnetic separation whereby the particles are suspended in a liquid
• CPC Class Codes: B03C 2201/26 for use in medical or biological applications
• CPC Class Codes: B03C 5/005 Dielectrophoresis, i.e. dielectric particles migrating towards the region of highest field strength
• CPC Class Codes: C12M 47/12 Purification C12M47/04 takes precedence
• 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 2710/00051 Methods of production or purification of viral material
• CPC Class Codes: C12N 7/00 Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00
• CPC Class Codes: C12N 7/02 Recovery or purification
• CPC Class Codes: C12Q 1/02 involving viable microorganisms
• CPC Class Codes: C12Q 1/24 Methods of sampling, or inoculating or spreading a sample; Methods of physically isolating an intact microorganisms
• CPC Class Codes: G01N 1/34 Purifying; Cleaning processes or apparatus for extracting or separating nucleic acids from biological samples C12N15/1003
• CPC Class Codes: G01N 33/491 by separating the blood components G01N15/05 takes precedence
• CPC Class Codes: G02B 21/361 Optical details, e.g. image relay to the camera or image sensor G02B21/364 takes precedence; illumination details G02B21/06 and subgroups

Abstract

Methods and systems for purifying cells and/or viruses are provided. The sample is added to a well disposed in a medium. A potential is applied across the medium to cause the contaminants to enter one or more walls of the well, and retain the cells and/or viruses in the well. The cells and/or viruses can be removed from the well, and optionally adhered or fixed to a surface, or detected. In one embodiment, the cells and/or viruses may be retained in the well by embedding in the medium. The medium including the embedded cells and/or viruses may be excised or otherwise removed and transferred to a glass slide or other solid surface.

Claims

1. A method of quantifying growth of one or more purified microorganisms, comprising:

adding a sample containing contaminants and one or more microorganisms to a well disposed in a porous medium;

applying an electrical potential across the medium to cause the contaminants to enter the porous medium through one or more walls of the well while retaining the one or more microorganisms in the well;

reversing the electrical potential for a sufficient time to displace the one or more microorganisms from a surface of the one or more walls of the well in the porous medium, thereby purifying the one or more microorganisms;

introducing the one or more purified microorganisms to a flow cell;

embedding the one or more purified microorganisms in an embedding medium, thereby immobilizing the one or more microorganisms in the flow cell;

imaging the one or more purified microorganisms using automated digital microscopy; and

quantifying growth of the one or more purified microorganisms over a period of time using imaging data.

2. The method of claim 1, wherein applying the electrical potential and reversing the electrical potential are performed iteratively.

3. The method of claim 1, wherein the one or more purified microorganisms are bacteria.

4. The method of claim 1, wherein the embedding medium comprises agar.

5. The method of claim 3, wherein the embedding medium comprises agar.

6. The method of claim 4, wherein the sample is mixed at least one of before applying the electrical potential, during application of the electrical potential, and after applying the electrical potential.

7. The method of claim 1, wherein the sample contains between 0.1 and 1×

10⁸ units/mL of microorganisms.

8. The method of claim 3, wherein the sample contains between 0.1 and 1×

10⁸ units/mL of microorganisms.

9. The method of claim 1, wherein the sample comprises a blood sample.

10. The method of claim 1, wherein the sample comprises a lung sample.

11. The method of claim 1, wherein the sample comprises a urine sample.

12. The method of claim 1, wherein the porous medium comprises a hydrogel.