Method and apparatus for selective removal of cells from a cell suspension by mechanical lysis

Patent metadata

Field	Value
Publication number	US-12072333-B2
Application number	US-201716346562-A
Country	US
Kind code	B2
Filing date	Nov 2, 2017
Priority date	Nov 2, 2016
Publication date	Aug 27, 2024
Grant date	Aug 27, 2024

Abstract

In some embodiments, the present disclosure provides an apparatus and methods for selectively lysing cells from a cell suspension including disassociating a population of cells from a tissue sample, the population of cells including a population of one or more viable cells and a population of one or more dead and dying cells, exposing the population of cells to at least one pulse of fluid shear stress having a force along a conduit wall of from about 500 dyn/cm2 to about 2500 dyn/cm2 to substantially lyse the population of one or more dead or dying cells, leaving the population of one or more viable cells substantially intact, and collecting the population of one or more viable cells.

First claim

What is claimed is:

1. A method comprising:
disassociating a population of cells from a tissue sample, the population of cells including a population of one or more viable cells and a population of one or more dead and dying cells;
exposing the population of cells to at least one pulse of fluid shear stress having a force along a conduit wall of from about 500 dyn/cm2 to about 2500 dyn/cm2 to substantially lyse the population of one or more dead or dying cells, leaving the population of one or more viable cells substantially intact; and
collecting the population of one or more viable cells.

2. The method of claim 1 , further comprising isolating at least one individual cell from the population of one or more viable cells.

3. The method according to claim 2 , wherein the isolation of at least one individual cell from the population of cells is accomplished in less than 4 hours.

4. The method of claim 2 , further comprising isolating a population of nucleic acid sequences from the at least one individual cell, wherein the population of nucleic acid sequences has not substantially changed from prior to disassociation.

5. The method of claim 2 , further comprising isolating a population of amino acid sequences from the at least one individual cell, wherein the population of amino acid sequences has not substantially changed from prior to disassociation.

6. The method of claim 2 , wherein the isolation of at least one individual cell from the population of cells is accomplished in less than 3.5 hours.

7. The method of claim 2 , wherein the at least one individual cell comprises transcribed RNA and wherein the transcribed RNA is characterized.

8. The method of claim 1 , wherein exposing the population of cells to the at least one pulse of fluid shear stress comprises at least one pulse having a duration of between about 1 millisecond and about 10 milliseconds.

9. The method of claim 1 , wherein the exposing the population of cells to the at least one pulse of fluid shear stress comprises exposing the population of cells to the at least one pulse of fluid shear stress having a force of between about 500 dyn/cm 2 and about 2,500 dyn/cm 2 with the force being measured along a conduit wall wherein the at least one pulse of fluid shear stress has a force of between about 850 dyn/cm 2 and about 1500 dyn/cm 2 .

10. The method of claim 1 , wherein collecting the population of one or more viable cells from the population of cells comprises substantially removing the lysed dead and dying cells by density gradient centrifugation, centrifugation, size-based filtration, cell sorters, or microfluidics.

11. The method of claim 1 , wherein the exposing the population of cells to the at least one pulse of fluid shear stress comprises exposing the population of cells to the at least one pulse of fluid shear stress having a force of between about 600 dyn/cm 2 and about 1,500 dyn/cm 2 with the force being measured along a conduit wall.

12. The method of claim 1 , wherein at least about 90% of the viable cells survive the fluid shear stress.

13. The method of claim 1 , wherein the population of cells from a tissue sample is nonhomogeneous.

14. The method of claim 1 , wherein a population of cells is exposed to at least two pulses of fluid shear stress and wherein the at least two pulses of fluid shear stress are substantially the same.

15. The method of claim 1 , further comprising collecting the lysed dead and dying cells.

16. The method of claim 15 , further comprising characterizing the nucleic acid sequences or amino acid sequences in the collected lysed dead and dying cells.

Assignees

• Univ Iowa Res Found
• Synderbio Inc

Inventors

• Henry Michael D
• Krog Benjamin
• Vigmostad Sarah
• Cable Mike

Classifications

• G01N1/34 Primary

  Purifying; Cleaning {(processes or apparatus for extracting or separating nucleic acids from biological samples C12N15/1003)} · CPC title

• G01N33/4833

  of solid biological material, e.g. tissue samples, cell cultures (tissue in vivo A61B5/00; cell suspensions G01N33/48735) · CPC title

• G01N2001/2866

  Grinding or homogeneising · CPC title

• G01N1/286

  involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising (microtomes G01N1/06; pulverising in general B02C; mixing in general B01F) · CPC title

• C12Q1/6806

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title