# US 10,768,141 B2 · Isoelectric focusing arrays and methods of use thereof > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,768,141 B2 - Document ID: US10768141B2 - Status: Granted - Kind: B2 - Application number: 15/749,039 - Filed: 2016-09-08 - Published: 2018-08-02 - Granted: 2020-09-08 - Earliest claimed: 2015-09-11 - Assignees: The Regents of the University of California - Applicants: The Regents of the University of California - Inventors: Amy E. Herr; Augusto M. Tentori; Kevin A. Yamauchi - Web page: https://patentgrep.com/patent/US10768141B2 ## CPC - G01N 27/44747: {Composition of gel or of carrier mixture} - B01D 57/02: by electrophoresis (treatment of water, waste water, sewage or sludge by electrophoresis C02F1/469; electrophoretic production of compounds or non-metals C25B7/00; investigating or analysing materials by using electrophoresis G01N27/26) - G01N 27/44791: {Microapparatus (sample containers with integrated microfluidic structures B01L3/5027)} - G01N 27/44795: {Isoelectric focusing} - 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) - 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) ## Abstract Isoelectric focusing devices configured for multiplex separation of sample components of interest in a polymeric separation medium are provided. Also provided are methods of using the devices as well as systems and kits that include the devices. The devices, systems and methods find use in a variety of different applications, including diagnostic and validation assays. ## Claims ### Claim 1 A device comprising: a polymeric separation medium comprising two or more isoelectric focusing regions each comprising an acidic boundary sub-region, a basic boundary sub-region, and a focusing sub-region positioned between the acidic boundary sub-region and the basic boundary sub-region, wherein each isoelectric focusing region is configured for multiplex separation of sample components of interest from two or more samples in the polymeric separation medium, wherein the two or more isoelectric focusing regions are arranged in series along an axis of the polymeric separation medium, and wherein each isoelectric focusing region comprises a pH gradient along the axis of the polymeric separation medium. ### Claim 2 (depends on 1) The device of claim 1, wherein the isoelectric focusing region comprises a polymeric gel. ### Claim 3 (depends on 2) The device of claim 2, wherein the pH gradient is an immobilized pH gradient. ### Claim 4 (depends on 1) The device of claim 1, wherein the polymeric separation medium further comprises functional groups that covalently bond to the sample components of interest in the polymeric separation medium upon application of an applied stimulus. ### Claim 5 (depends on 1) The device of claim 1, wherein the polymeric separation medium comprises: a plurality of microwells arranged for parallel separations of the sample components of interest along the axis from each microwell. ### Claim 6 (depends on 5) The device of claim 5, wherein each microwell is dimensioned to accommodate a single cell. ### Claim 7 (depends on 5) The device of claim 5, further comprising a polymeric gel cover configured to contact a top surface of the polymeric separation medium. ### Claim 8 (depends on 7) The device of claim 7, wherein the polymeric gel cover contacts one or more electrodes configured to apply an electric field to the polymeric separation medium. ### Claim 9 (depends on 7) The device of claim 7, wherein the polymeric gel cover comprises a pH gradient along an axis of the polymeric gel cover substantially parallel to the axis of the polymeric separation medium. ### Claim 10 (depends on 7) The device of claim 7, wherein the polymeric gel cover comprises a lysis reagent. ### Claim 11 (depends on 5) The device of claim 5, wherein the polymeric separation medium comprises two or more isoelectric focusing regions arranged in series along the axis of the polymeric separation medium. ### Claim 12 (depends on 5) The device of claim 5, wherein the polymeric separation medium further comprises functional groups that covalently bond to the sample components of interest in the polymeric separation medium upon application of an applied stimulus. ### Claim 13 (depends on 1) A method comprising: contacting a sample to the polymeric separation medium of the device according to claim 1; and applying an electric field to the polymeric separation medium in a manner sufficient to perform a multiplex isoelectric focusing separation of the sample components of interest in the polymeric separation medium. ### Claim 14 (depends on 13) The method of claim 13, wherein the polymeric separation medium comprises two or more isoelectric focusing regions arranged in series along an axis of the polymeric separation medium. ### Claim 15 (depends on 13) The method of claim 13, wherein the polymeric separation medium comprises: a pH gradient along an axis of the polymeric separation medium; and a plurality of microwells arranged for parallel separations of the sample components of interest along the axis from each microwell; and wherein, prior to applying the electric field, the method comprises contacting a top surface of the polymeric separation medium with a polymeric gel cover comprising a pH gradient along an axis of the polymeric gel cover substantially parallel to the axis of the polymeric separation medium. ### Claim 16 (depends on 15) The method of claim 15, wherein the sample comprises cells and/or cellular components. ### Claim 17 (depends on 16) The method of claim 16, further comprising lysing the cells to produce cellular components of interest in the sample. ### Claim 18 (depends on 13) The method of claim 13, further comprising immobilizing the separated sample components in the polymeric separation medium. ### Claim 19 (depends on 13) The method of claim 13, further comprising detecting the separated sample components. ### Claim 20 (depends on 1) A system comprising: the device according to claim 1; and an electric field source. ### Claim 21 (depends on 1) A kit comprising: the device according to claim 1; and a packaging containing the device. ### Claim 22 (depends on 1) The device of claim 1, wherein the acidic boundary sub-region comprises immobilines that buffer at an acidic pH and the basic boundary sub-region comprises immobilines that buffer at a basic pH. ### Claim 23 (depends on 1) The device of claim 1, wherein the acidic boundary sub-region is positioned on a same side of the focusing sub-region as a positive electrode, and the basic boundary sub-region is positioned on a same side of the focusing sub-region as a negative electrode.