METHOD FOR SEPARATING CELLS-BEAD COMPLEXES

• US 20150253226A1
• Filed: 09/20/2013
• Published: 09/10/2015
• Est. Priority Date: 09/21/2012
• Status: Abandoned Application

• Current Assignee: AcouSort AB
• Original Assignee: AcouSort AB
• Inventors: Augustsson, Per, Laurell, Thomas, Lenshof, Andreas, Scheding, Stefan
• Application Number:
  US14/430,033
• Publication Number:
  US 20150253226A1
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: B01L 2200/0652 Sorting or classification of particles or molecules
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2400/0436 acoustic forces, e.g. surface acoustic waves [SAW]
• CPC Class Codes: B01L 3/502753 characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation separation in general B01D; microapparatus for analysis using electrophoresis G01N27/44791; sample preparation G01N1/28
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: B01L 3/502776 specially adapted for focusing or laminating flows
• CPC Class Codes: G01N 1/4077 by other techniques involving separation of suspended solids
• CPC Class Codes: G01N 2001/4094 using ultrasound
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form
• CPC Class Codes: G01N 33/54373 involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G01N 33/569 for microorganisms, e.g. protozoa, bacteria, viruses
• CPC Class Codes: G01N 33/56972 White blood cells

Abstract

The invention relates to a method and system to separate a subgroup of cells from a mixture of cells. This is achieved by creating a complex consisting of the subgroup of cells that are to be separated and a ligand that could be constructed of molecules such as antibodies, monoclonal antibodies, aptamers or fragments thereof specific for said subgroup of cells and microbeads present in a suspension. The suspension is then exposed to an acoustic field in one or two dimensions, which forces said cells and cell-bead complexes to separate from each other based on differences in size, density and compressibility.

Claims

1. A micro scale method for separating cell-bead complexes comprising cells, ligands and beads from a mixture of different types of cells in a suspension, comprising the steps of;

i) subjecting a suspension to pressure, wherein said pressure forces said suspension into at least one inlet and into at least one pre-alignment channel present on a microfluidic chip,

ii) subjecting said suspension to a two dimensional acoustic force directed perpendicular to the length direction of the pre-alignment channel,

iii) forcing said mixture of cells cell-bead complexes through the side branches of a trifurcated inlet while forcing cell free ASGL through the central branch of said trifurcated inlet simultaneously into at least one separation channel, wherein said mixture of cells and cell-bead complexes are pushed by hydrodynamic forces towards the walls of the channel,

iv) subjecting said cells and complexes to a one dimensional acoustic force directed perpendicular to the length direction towards the center of the separation channel and

v) collecting cell-bead complexes at the end of the separation channel present on the microfluidic chip through at least one outlet.

vi) collecting non-complexed cells at the end of the separation channel present on the microfluidic chip through at least one outlet.

2. The method according to claim 1, wherein said ligand is antibodies, monoclonal antibodies, affibodies, aptamers, single chain antibodies, antigen specific molecules or fragments thereof.

3. The method according to claim 2, wherein said cells are selected from the group consisting erythrocytes, platelets or leukocytes dendritic cells, stem cells/precursor cells, endothelial cells and epithelia cells.

4. The method according to claim 1, wherein said beads are selected from the group consisting of the beads may be polymer beads, superparamagnetic beads, silica beads, metal beads.

5. The method according to claim 1, wherein said ASGL has a density between 0.900-1.100 g/cm³.

6. The method according to claim 1, wherein the two dimensional acoustic force in step ii) consists of ½

wavelength and 1 wavelength perpendicular to each other.

7. The method according to claim 1, wherein said one dimensional acoustic force in step iv) consists of ½

wavelength.

8. The method according to claim 1, wherein said two dimensional acoustic force is caused by a frequency of from 1 to 10 MHz.

9. The method according to claim 1, wherein said method is maintained at a constant temperature.

10. The method according to claim 1, wherein the height or the width of channel is from 75 um to 800 um.