Filter Method for Separating Unbound Ferrofluid from Target-bound Ferrofluid in a Biological Sample

• US 20120055854A1
• Filed: 02/05/2010
• Published: 03/08/2012
• Est. Priority Date: 02/05/2009
• Status: Abandoned Application

• Current Assignee: Tibbe, Arjan G.J.
• Original Assignee: Tibbe, Arjan G.J.
• Inventors: Tibbe, Arjan G.J.
• Application Number:
  US13/147,908
• Publication Number:
  US 20120055854A1
• Time in Patent Office: Days
• US Class Current: 209/214
• CPC Class Codes: B03C 1/01 by addition of magnetic adjuvants
• CPC Class Codes: B03C 1/286 disposed at the inner circumference of a recipient, e.g. magnetic drain bolt
• CPC Class Codes: B03C 2201/18 Magnetic separation whereby the particles are suspended in a liquid

Abstract

A filtering system for separating unbound ferrofluid from bound ferrofluid in the enrichment of a target entity in a biological sample. The filtering device of the present invention has application in the isolation target cells from unbound ferrofluid during separation with a permanent magnet (floater) mechanism. This process reduces interference of unbound ferrofluid during subsequent image analysis or enumeration of cells by image cytometry. The system has application in the assessment of target populations such as leukocyte subsets in different bodily fluids or bacterial contamination in environmental samples, food products and bodily fluids. Briefly, fluorescently labeled target cells are linked to magnetic particles or beads. The linkage process results in a mixture having a population of contaminating unbound magnetic particles. In one embodiment for separation, a small, permanent magnet is inserted directly into the chamber containing the labeled cells. The magnets are coated with PDMS silicone rubber to provide a smooth and even surface which allows imaging on a single focal plane. A filter is positioned on a cover of the floater device to allow unbound ferrofluid to pass through the pores, but restrict the passage of the target entity. The floater and filter are removed from the sample and the filter surface is illuminated with fluorescent light emitted by the target cells captured by a CCD camera. Image analysis can be performed with a novel algorithm to provide a count of the cells on the surface, reflecting the target cell concentration of the original sample.

Claims

1. A method for separating unbound immunomagnetic particles from target bound immunomagnetic particles in a biological specimen, comprising:

a. obtaining said biological specimen from a subject, wherein said specimen contains a mixture of unbound immunomagnetic particles and immunomagnetic particles bound to a target population;

c. adding a small permanent magnet directly to said specimen wherein the collection surface on said magnet is separated from said target population by a filter with a porosity size between said unbound immunomagnetic particles and said target population; and

d. separating said target population from said unbound immunomagnetic particles wherein said target population is collected on the surface of said filter and said unbound ferrofluid is collected on the collection surface of said magnet.

2. The target population of claim 1 wherein the target population are cells.

3. The target population of claim 2 wherein said target population is CD4 expressing cells.

4. The small permanent magnet of claim 1 wherein said magnet is neodymium.

5. The small permanent magnet of claim 1 wherein said magnet is a disc having a diameter of about 1.6 mm and a height of 0.8 mm.

6. The small permanent magnet of claim 1 coated with PDMS silicone.

7. The filter of claim 1 wherein said filter is a pillar structure.

8. A device for separating unbound immunomagnetic particles from target bound immunomagnetic particles in a biological specimen, comprising:

a. a filter with a porosity size between said unbound immunomagnetic particles and said target population;

b. a magnet with a collection surface; and

c. a spacer means to separate said filter from said collection surface wherein said spacer provides a volume to allow unbound immunomagnetic particles to traverse the filter and collect on said collection surface.

9. The filter of claim 8 wherein said filter is a pillar structure.