# US 8,148,169 B2 · Method and device for extracting an analyte

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- Number: US 8,148,169 B2
- Document ID: US8148169B2
- Status: Granted
- Kind: B2
- Application number: 13/091,634
- Filed: 2011-04-21
- Published: 2011-08-11
- Granted: 2012-04-03
- Earliest claimed: 2004-05-12
- Assignees: PhyNexus, Inc.
- Inventors: Douglas T. Gjerde; Ronald Jones; Allen Burge
- Web page: https://patentgrep.com/patent/US8148169B2

## CPC

- B01J 20/285: based on polymers
- B01D 15/1896: {Membrane chromatography or membrane adsorbers}
- B01D 15/3804: {Affinity chromatography}
- B01J 20/291: Gel sorbents
- B01L 3/0275: {Interchangeable or disposable dispensing tips}
- B82Y 30/00: Nanotechnology for materials or surface science, e.g. nanocomposites
- B01D 15/325: {Reversed phase}
- B01D 15/327: {with hydrophobic interaction}
- B01J 2220/64: In a syringe, pipette, e.g. tip or in a tube, e.g. test-tube or u-shape tube
- B01L 2200/0631: Purification arrangements, e.g. solid phase extraction [SPE]
- G01N 2030/009: {Extraction}
- G01N 2035/1053: {for separating part of the liquid, e.g. filters, extraction phase}
- Y10T 436/25375: Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
- Y10T 436/255: including use of a solid sorbent, semipermeable membrane, or liquid extraction
- Y10T 436/2575: Volumetric liquid transfer
- B01J: CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01D: SEPARATION (separating solids from solids by wet methods B03B, B03D; by pneumatic jigs or tables B03B; by other dry methods B07; magnetic or electrostatic separation of solid materials from solid materials or fluids, separation by high-voltage electric fields B03C; centrifuges, vortex apparatus B04; presses per se for squeezing-out liquid from liquid-containing material B30B9/02; treatment of water C02F, e.g. softening by ion-exchange C02F1/42; {arrangements of air intake cleaners in gas turbine plants F02C7/05} ; arrangements or mounting of filters in air-conditioning, air-humidification or ventilation F24F13/28)
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B82Y: SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION

## 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 method involves adjusting the head pressure of the column during the process, or otherwise controlling or regulating the head pressure.

## Claims

### Claim 1

An automated parallel method for passing liquid through a plurality of pipette tip columns comprising the steps of:
a) providing a plurality of pipette tip columns, wherein each column is comprised of:
i. a column body having an open upper end for communication with a pump, a first open lower end for the uptake and dispensing of fluid, and an open passageway between the upper and lower ends of the column body,
ii. a bottom frit, wherein the bottom frit is at or near the open lower end of the pipette tip column,
iii. an optional top frit positioned between the bottom frit and the open upper end of the column body,
iv. a bed of hydrated gel resin media,
v. a storage liquid, wherein the storage liquid is comprised of a water miscible solvent having a viscosity greater than that of water, and
vi. a pump sealingly attached to the open upper end of the column, wherein the pump is comprised of a piston, and wherein actuation of the pump piston causes liquid to be drawn into or expelled from the bed of hydrated gel resin media;
b) contacting said open lower ends of the columns with a liquid;
c) actuating the pump piston to draw the liquid into the open lower ends of the columns and through the bed of hydrated gel resin media, wherein the volume of liquid drawn into the columns is 250 μl or less and wherein the pump piston displacement is 20 to 75 μl greater than the volume of liquid drawn into the column; and
d) actuating the pump piston to expel the liquid through the bed of hydrated gel resin media and out of the open lower end of the columns.

### Claim 2 (depends on 1)

The method of claim 1, wherein the water miscible solvent has a boiling point greater than 150° C.

### Claim 3 (depends on 2)

The method of claim 2, wherein said water miscible solvent comprises 50% of the storage liquid.

### Claim 4 (depends on 3)

The method of claim 3, wherein the water miscible solvent comprises a diol, triol, or polyethylene glycol of n=2 to n=150.

### Claim 5 (depends on 1)

The method of claim 1, wherein the water miscible solvent is glycerol.

### Claim 6

An automated parallel method for passing liquid through a plurality of pipette tip columns comprising the steps of:
a) providing a plurality of pipette tip columns, wherein each column is comprised of:
i. a column body having an open upper end for communication with a pump, a first open lower end for the uptake and dispensing of fluid, and an open passageway between the upper and lower ends of the column body,
ii. a bottom frit, wherein the bottom frit is at or near the open lower end of the pipette tip column,
iii. an optional top frit positioned between the bottom frit and the open upper end of the column body,
iv. a bed of resin media, and
v. a pump sealingly attached to the open upper end of the column, wherein the pump is comprised of a piston, and wherein actuation of the pump piston causes liquid to be drawn into or expelled from the bed of resin media;
b) contacting said open lower ends of the columns with a liquid;
c) actuating the pump piston to draw the liquid into the open lower ends of the columns and through the bed of resin media, wherein the volume of liquid drawn into the columns is 250 μl or less wherein the pump piston displacement is 20 to 75 μl greater than the volume of liquid drawn into the column; and
d) actuating the pump piston to expel the liquid through the bed of resin media and out of the open lower end of the columns.

### Claim 7 (depends on 6)

The method of claim 6, wherein the bed resin media is agarose or sepharose.

### Claim 8 (depends on 6)

The method of claim 6, wherein the bed of resin media is further comprised of an affinity binding agent.

### Claim 9 (depends on 6)

The method of claim 6, wherein the analyte is a protein.

### Claim 10 (depends on 6)

The method of claim 6, wherein the method is performed simultaneously on at least 8 and at most 96 pipette tip columns.