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Tandem paired column chemistry for high-throughput proteomic exosome analysis

An analytical column, protease technology, applied in the fields of omics, organic chemistry, analytical materials, etc., can solve the problems of time-consuming, insufficient target peptide/protein sensitivity, purity and sensitivity problems, etc.

Active Publication Date: 2022-08-02
NX PRENATAL INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, even with samples obtained from extracellular vesicles, issues of purity and sensitivity remain
[0020] In summary, current proteomic analysis methods suffer from insufficient sensitivity and imprecision in the quantification of target peptides / proteins due to the above reasons
Additionally, current methods are time-consuming and have low yields

Method used

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  • Tandem paired column chemistry for high-throughput proteomic exosome analysis
  • Tandem paired column chemistry for high-throughput proteomic exosome analysis
  • Tandem paired column chemistry for high-throughput proteomic exosome analysis

Examples

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Embodiment

[0113] Frozen maternal plasma samples were used. Samples can be shipped on dry ice and stored at -80°C.

[0114] Microparticles were enriched by size exclusion chromatography (SEC) and eluted isocratically with water (RNase-free, DNase-free distilled water). Size exclusion chromatography procedures were performed using columns manufactured by AmericanBio (Natick, MA). Briefly, these columns were loaded with 10 mL of 2% Sepharose 2B (60-200 um pore size) purchased from GE Healthcare Life Sciences (Marlborough, MA). The columns were stored under refrigerated conditions. They were allowed to equilibrate to room temperature before using the columns for exosome isolation. After equilibration to room temperature, the column was washed with RNase-free, DNase-free distilled water. At least 0.5 mL of thawed purified plasma was applied to the column. Microparticles were eluted with RNase-free, DNase-free distilled water.

[0115] Circulating particles are trapped in the column voi...

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Abstract

The present application provides compositions and methods for sample preparation and mass spectrometry analysis of peptide samples obtained from biological samples. The compositions and methods include in-line column systems in which a trapping column is in fluid contact with an analytical column such as, for example, an HPLC column. As the analytes are eluted from the analytical column, they can be passed to a detector (eg, a mass spectrometer) for peptide analysis.

Description

[0001] Statement Regarding Federally Funded Research [0002] none. [0003] Citations to Related Applications [0004] This application claims the benefit of priority from US Application No. 62 / 757,922 (Rosenblatt et al.), filed on November 9, 2018, the entire contents of which are incorporated herein in their entirety. [0005] The names of the parties to the joint research agreement [0006] none. [0007] sequence listing [0008] none. technical field [0009] The present application is in the field of sample preparation for peptide analysis, eg, analysis by high performance liquid chromatography and / or mass spectrometry. Background technique [0010] Many diagnostic procedures rely on proteomic analysis. The proteome, the unique collection of proteins present in specific cells in specific developmental states, can provide a wealth of information about cellular conditions and has found use in diagnostic and therapeutic strategies, especially for cancer and pregnan...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01D3/14B01D15/08B01L3/00C07K1/16G01N30/02G01N33/48H01J49/26
CPCB01D15/325B01D15/34G01N33/6851G01N33/689B01L3/502715B01L2400/0622B01L3/502753B01L2400/0487G01N30/08G01N2030/085G01N2030/8831G01N33/5076G01N33/6848G01N1/4044G01N1/405G01N30/88G01N2570/00
Inventor K·P·罗森布拉特N·哈贾M·哈立德
Owner NX PRENATAL INC
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