Method and device for extracting an analyte

• US 7,488,603 B2
• Filed: 08/17/2004
• Issued: 02/10/2009
• Est. Priority Date: 07/14/2003
• Status: Active Grant

• Current Assignee: PhyNexus, Inc.
• Original Assignee: PhyNexus, Inc.
• Inventors: Gjerde, Douglas T., Hanna, Christopher P.
• Primary Examiner(s): Warden; Jill
• Assistant Examiner(s): JARRETT, LORE RAMILLANO
• Application Number:
  US10/920,960
• Publication Number:
  US 20050016921A1
• Time in Patent Office: 1,638 Days
• Field of Search: 422/101, 436/66, 436/177, 210/198.2
• US Class Current: 436/177
• CPC Class Codes: B01D 15/325 Reversed phase
• CPC Class Codes: B01D 15/327 with hydrophobic interaction
• CPC Class Codes: B01D 15/3804 Affinity chromatography
• CPC Class Codes: B01J 20/285 based on polymers
• CPC Class Codes: B01J 20/291 Gel sorbents
• CPC Class Codes: B01J 2220/64 In a syringe, pipette, e.g. tip or in a tube, e.g. test-tube or u-shape tube
• CPC Class Codes: B01L 2200/0631 Purification arrangements, e.g. solid phase extraction [SPE]
• CPC Class Codes: B01L 3/0275 Interchangeable or disposable dispensing tips
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: G01N 2030/009 Extraction
• CPC Class Codes: G01N 2030/525 surface properties, e.g. porosity
• CPC Class Codes: G01N 2035/103 using disposable tips
• CPC Class Codes: G01N 2035/1039 Micropipettes, e.g. microcapillary tubes
• CPC Class Codes: G01N 2035/1053 for separating part of the liquid, e.g. filters, extraction phase
• CPC Class Codes: G01N 30/6004 end pieces
• CPC Class Codes: G01N 35/10 Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices G01N35/0099 takes precedence
• CPC Class Codes: Y10T 436/153333 Halogen containing
• CPC Class Codes: Y10T 436/25375 Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
• CPC Class Codes: Y10T 436/255 including use of a solid sorbent, semipermeable membrane, or liquid extraction
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

The invention provides extraction columns for the purification of an analyte (e.g., a biological macromolecule, such as a peptide, protein or nucleic acid) from a sample solution, as well as methods for making and using such columns. The columns typically include a bed of extraction media positioned in the column between two frits. In some embodiments, the extraction columns employ modified pipette tips as column bodies. In some embodiments, the invention provides methods characterized by the elution of analyte in a small volume of liquid.

Claims

1. A method of extracting an analyte from a solution comprising the steps of:

a. passing a solution containing an analyte back and forth through a pipette tip column, wherein the pipette tip column is comprised of;

i. a column body having an open upper end, an open lower end, and an open channel between the upper and lower ends of the column body,

ii. a bottom frit bonded to and extending across the open channel,

iii. a top frit bonded to and extending across the open channel between the bottom frit and the open upper end of the column body, wherein the top frit, bottom frit, and column body define the extraction media chamber, wherein the top frit and the bottom frit are less than 350 microns thick, wherein the bottom frit is a membrane screen and the top frit is optionally a membrane screen, and wherein the membrane screen is selected from a group of materials consisting of nylon, polyester, polyamide, polycarbonate, cellulose, polyethylene, nitrocellulose, cellulose acetate, polypropylene, polysulfone, and glass, and

iv. a bed of extraction media having a bed volume, whereby analyte adsorbs to said extraction media; and

b. eluting the analyte by aspirating and discharging a desorption solvent through the pipette tip column, and wherein said analyte is an active protein.

2. The method of claim 1, wherein the volume of desorption solvent is in the range of about 10% to about 300% of the bed volume.

3. The method of claim 1, wherein step (a) is performed more than one time.

4. The method of claim 1, wherein the bed volume is in the range of between 0.1 and 100 uL.

5. The method of claim 1, wherein between step (a) and step (b), a quantity of wash solution is aspirated and discharged through the extraction media.

6. The method of claim 1, wherein step (b) is performed more than one time.

7. The method of claim 1, wherein the extraction media comprises an affinity binding agent.

8. The method of claim 1, wherein the bed of extraction media comprises a packed bed of gel resin beads.

9. the method of claim 8, wherein the gel resin beads are agarose beads or sepharose beads.

10. The method of claim 1, wherein the bottom frit has a pore volume of 1 microliter or less.

11. The method of claim 1, wherein the bottom frit is less than 200 microns thick.

12. The method of claim 1, wherein the upper end of the column body is attached to a pump for aspirating fluid through the lower end of the column body.

13. The method of claim 12, wherein the pump is a pipettor, a syringe, or a peristaltic pump.