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Tunable electroosmotic flow polymer coated capillary

A polymer and capillary technology, applied in the field of tunable electroosmotic flow polymer-coated capillary, can solve the problem that the operating performance needs to be improved

Active Publication Date: 2020-03-17
UNIV OF NOTRE DAME DU LAC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Finally, RAFT-polymerized coated columns were not evaluated for proteomic applications in CZE
[0010] There is a problem with polymer-coated capillaries for separation of proteins or protein digests by CZE.

Method used

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  • Tunable electroosmotic flow polymer coated capillary
  • Tunable electroosmotic flow polymer coated capillary
  • Tunable electroosmotic flow polymer coated capillary

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0137] Example 1. Low electroosmotic flow polymer coating

[0138] Reagents and chemical supplies. Formic acid (FA), acetic acid (HAc), acrylamide, [2-(methacryloyloxy)ethyl] trimethylammonium chloride solution (80wt.% in H 2 O), 4,4-azobis(4-cyanovaleric acid), cyanomethyl[3-(trimethoxysilyl)propyl]trithiocarbonate, ammonium acetate, TPCK treated Bovine pancreatic trypsin, benzyl alcohol, dithiothreitol (DTT) and iodoacetamide (IAA) were purchased from Sigma-Aldrich (St. Louis, USA). Pierce TM HeLa protein digestion standards were purchased from Thermo Fisher Scientific (Hanover Park, USA). Methanol was purchased from Honeywell Burdick & Jackson (Wicklow, Ireland). Uncoated fused silica capillaries (50 μm inner diameter x 150 μm outer diameter) were purchased from PolymicroTechnologies (Phoenix, AZ). The water deionization process adopts the Nano Pure system of Thermo Scientific (Marietta, OH).

[0139] The positively charged capillary was prepared by the SCARAFT method. Usi...

Embodiment 2

[0151] Example 2. Sample preparation

[0152] Prepare E. coli samples. Solid lysogenic broth (LB) was used to prepare agar plates for E. coli culture. Solid LB was prepared by dissolving 3 g NaCl, 3 g trypsin, 1.5 g yeast extract, and 6 g agar in 300 mL deionized water. A liquid LB medium (without agar) for E. coli culture was also prepared by mixing 10 g NaCl, 10 g trypsin p, and 5 g yeast extract in 1 L of deionized water. Before use, autoclave all media, plates, other utensils and flasks. The frozen culture of E. coli (Dh5-Alpha) was thawed and plated on the prepared agar plate. After incubating for 24 hours at 37°C, a single colony was isolated and grown in a test tube containing 4 mL of liquid LB medium, and incubated overnight in a shaker at 37°C. When the contents of the test tube become opaque, transfer the liquid culture medium to a new flask and shake it overnight at 37°C. The liquid LB medium with Escherichia coli was centrifuged, and the obtained Escherichia coli...

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Abstract

A surface-confined aqueous reversible addition-fragmentation chain transfer (SCARAFT) polymerization method was developed to coat capillaries for use in capillary zone electrophoresis (CZE). This coating produced an electroosmotic an order of magnitude lower than that of commercial linear polyacrylamide (LPA)-coated capillaries. Coated 5 capillaries were evaluated for bottom-up proteomic analysisusing CZE. The very low electroosmotic mobility results in a 200 min separation and improved single-shot analysis. Various types of coatings were prepared by simply changing the functional vinyl monomers in the polymerization mixture.

Description

[0001] Related application [0002] This application, according to U.S.C. §119(e), claims the priority of the following earlier application: U.S. Provisional Patent Application 62 / 488,892 filed on April 24, 2017, which is hereby incorporated by reference. [0003] Government funding [0004] The present invention received government funding provided by the National Institutes of Health with the project number R01GM096767. The US government has certain rights in this invention. Background technique [0005] Capillary zone electrophoresis (CZE), due to its high sensitivity, orthogonality to reversed-phase chromatography and low cost, has attracted more and more attention from mass spectrometry-based proteomics. Although the separation is based on the charge / size ratio of the analyte in CZE, the characteristics of the capillary play an important role in electrophoresis performance. The non-specific adsorption of the analyte to the silanol group on the inner surface of the capillary can...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01D57/02B05D5/12C07K1/26
CPCB01D57/02C07K1/26G01N27/44752C08L53/005C09D133/16G01N27/44747
Inventor 诺曼·多维奇张振斌
Owner UNIV OF NOTRE DAME DU LAC