Kit for density-based separation of biological analytes using multiphase systems

Patent metadata

Field	Value
Publication number	US-9857353-B2
Application number	US-201514834593-A
Country	US
Kind code	B2
Filing date	Aug 25, 2015
Priority date	Aug 20, 2010
Publication date	Jan 2, 2018
Grant date	Jan 2, 2018

Abstract

A kit for separating a sample comprising one or more biological analytes of interest using a multi-phase system comprising: a) two or more phase components selected from the group consisting of a polymer, a surfactant, and combinations thereof; b) optionally a tag molecule capable of binding the one or more biological analytes of interest; and c) instructions for: (i) combining the two or more phase-separated solutions with a common solvent to create a multi-phase system; (ii) optionally, combining the biological analyte of interest and tag molecule, and (iii) separating the biological analyte of interest from the sample.

First claim

What is claimed is:

1. A kit for density based separating a sample comprising one or more biological analytes of interest using a multi-phase system having a common solvent; wherein each analyte has at least one dimension greater than 200 nm and is independently selected from the group consisting of protein, saccharide, polyterpene, polynucleic acid, cell, tissue, cell organelle, cell membrane, cell membrane fragments, virus, virus-like particle and parasite; and the kit comprises:
a) two or more different phase components comprising at least a first and second phase components which are both polymers; wherein when combined, the two or more phase components and the common solvent are selected to form a multi-phase system comprising phase-separated first and second phase solutions comprising the first and second phase components, respectively; and wherein the first and second phase components and the common solvent are selected such that the analyte is insoluble in the multi-phase system and interaction between the analyte and the multi-phase system is predominantly gravity-driven;
b) optionally a tag molecule capable of binding the one or more biological analytes of interest to form a tag molecule-analyte adduct having a density different from that of the biological analyte of interest;
c) a column, a well plate, a cuvette, a filament, a sheet, or a tube for supporting the multi-phase system; and
d) instructions for:
(i) combining the two or more phase components with the common solvent to create the multi-phase system;
(ii) optionally, combining the biological analyte of interest and tag molecule to form a tag molecule-analyte adduct,
(iii) introducing a sample comprising one or more biological analytes of interest or tag molecule-analyte adducts to the multi-phase system without disrupting the phase-separated solutions; and
(iv) separating the biological analyte of interest from the sample by allowing the biological analyte or the tag molecule-analyte adduct to migrate to a location in the multi-phase system characteristic of its density.

2. The kit of claim 1 , further comprising an aliquot of the common solvent which, when combined with the two or more phase components, provides a multiphase system.

3. The kit of claim 1 , wherein the instructions direct that the biological analyte of interest be combined with the tag molecule to form a tag molecule-analyte adduct before introduction to the multi-phase system.

4. The kit of claim 1 , wherein the instructions further direct that the biological analyte of interest and tag molecule be added to the multi-phase system to combine to form a tag molecule-analyte adduct in the multi-phase system.

5. The kit of claim 1 , wherein the kit further comprises one or more additives selected from the group consisting of dyes, nutrients, vitamins, antibiotics, anticoagulants, and buffers for combination with the phase components.

6. The kit of claim 1 , wherein the multi-phase system comprises one or more biologically compatible phases.

7. The kit of claim 1 , wherein the kit comprises a tag that has an affinity for one or more analytes of interest.

8. The kit of claim 1 , further comprising a lytic agent for introduction into one or more phases of the multiphase system.

9. The kit of claim 2 , wherein the common solvent is an aqueous solvent.

10. The kit of claim 2 , wherein the common solvent is an organic solvent.

11. The kit of claim 1 , wherein the gravity comprises enhanced gravitational force using centrifugation.

12. The kit of claim 1 , wherein the tube comprises a capillary tube, a test tube, a falcon tube, or a culture tube.

13. The kit of claim 1 , wherein the polymer is selected from the group consisting of dextran, polysucrose, poly(vinyl alcohol), poly(2-ethyl-2-oxazoline), poly(methacrylic acid), poly(ethylene glycol), polyacrylamide, polyethyleneimine, hydroxyethyl cellulose, polyvinylpyrrolidone, carboxy-polyacrylamide, poly(acrylic acid), poly(2-acrylamido-2-methyl-1-propanesulfonic acid), poly(diallyldimethyl ammonium chloride), poly(styrene sulfonic acid), polyallylamine, alginic acid, dextran sulfate, chondroitin sulfate A, diethylaminoethyl-dextran, poly(2-vinylpyridine-N-oxide), polydimethylsiloxane, and poly(propylene glycol).

14. The kit of claim 1 , wherein the polymer is independently selected from the group consisting of dextran, polysucrose, poly(vinyl alcohol), poly(2-ethyl-2-oxazoline), poly(ethylene glycol), polyacrylamide, hydroxyethyl cellulose, polyvinylpyrrolidone, carboxy-polyacrylamide, and poly(2-vinylpyridine-N-oxide).

15. The kit of claim 1 , wherein the polymer is selected from the group consisting of GRAS polymers.

16. The kit of claim 1 , wherein the polymer is selected from the group consisting of homopolymers, random copolymers, block copolymers, graft copolymers, ter-polymers, dendrimers, star polymers and combinations thereof.

17. The kit of claim 1 , wherein the polymer is linear, branched and/or cross-linked.

Assignees

• Harvard College

Inventors

• Mace Charles R
• Kumar Ashok A
• Wirth Dyann F
• Whitesides George M

Classifications

• G01N33/5759

  involving compounds localised on the membrane of tumour or cancer cells · CPC title

• B03B5/442

  composition of heavy media · CPC title

• G01N33/5375

  by changing the physical or chemical properties of the medium or immunochemicals, e.g. temperature, density, pH, partitioning · CPC title

• B03D3/00

  Differential sedimentation · CPC title

• B03B5/28

  by sink-float separation · CPC title