Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Two-dimensional Gel Electrophoresis01:22

Two-dimensional Gel Electrophoresis

Two-dimensional gel electrophoresis is a high-resolution protein separation method first introduced by O' Farrell and Klose in 1975. This method involves protein separation by two dimensions, mass and charge, making it more accurate than one-dimensional gel electrophoresis.
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as  cells...
High-Performance Liquid Chromatography: Introduction01:11

High-Performance Liquid Chromatography: Introduction

High-performance liquid chromatography(HPLC), formerly referred to as High-pressure liquid chromatography, is a powerful technique used to separate, identify, and quantify components in complex mixtures. The term "high pressure" refers to using high pressure to push the liquid mobile phase through the tightly packed columns.
In HPLC, two phases play a critical role in the separation process:
Types Of Column Chromatography01:29

Types Of Column Chromatography

The stability and compatibility of column material with samples are crucial for efficient purification in chromatographic techniques. Various operating parameters such as pH, temperature, or solvent affect the packing of the column material, thereby determining the purification efficiency. The choice of column material also plays an essential role in deciding the operating parameters and can be modified based on the proteins that need to be purified.
Gel Filtration Chromatography
When the...
High-Performance Liquid Chromatography: Instrumentation00:57

High-Performance Liquid Chromatography: Instrumentation

High-performance liquid chromatography, or HPLC, is an analytical technique that separates liquid samples under high pressures. An HPLC instrument consists of glass bottles for storing solvents called mobile phase reservoirs. HPLC-grade solvents are used to maintain high purity, and the dissolved gases are removed using a degasser, such as a vacuum pumping system or sparging with helium. The solvents are then pumped into the analytical column using a screw-driven syringe or reciprocating pumps.
Principles Of Column Chromatography01:13

Principles Of Column Chromatography

The chromatography technique was first invented in 1901 by Michael S. Tswett, a Russian botanist, to separate plant pigments using organic solvents. Further, in 1941, Archer John Porter Martin and R. L. M. Synge modified the technique by packing silica gel into a column. A mixture of amino acids was then separated on the packed column using chloroform and water mixture as the mobile phase. This was the first report on column chromatography. At present, column chromatography is a widely used...
High-Performance Liquid Chromatography: Elution Process01:05

High-Performance Liquid Chromatography: Elution Process

In High-Performance Liquid Chromatography (HPLC), the elution process is critical to the separation of analytes and the quality of chromatographic results. Elution describes how compounds move through the column and separate based on their interactions with the mobile and stationary phases. This process determines the resolution, peak shape, and retention times in the chromatogram, which are essential for identifying and quantifying components in complex mixtures. Understanding the elution...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Yeast Molecular Display for the Screening and Production of Immunological Proteins Using Nanobody as an Example.

Methods in molecular biology (Clifton, N.J.)·2026
Same author

Autonomous ribosome biogenesis in vitro.

Nature communications·2025
Same author

Bacterial removal using liposomes and an anionic adsorber.

Journal of bioscience and bioengineering·2024
Same author

In vitro selection of dye-fluorescence-enhancing peptide aptamer by cDNA display.

Analytical biochemistry·2024
Same author

Quantitative <sup>31</sup>P-NMR for the Purity Determination of the Organophosphorus Compound Brigatinib and Its Method Validation.

Chemical & pharmaceutical bulletin·2023
Same author

Secretion of mitochondrial DNA via exosomes promotes inflammation in Behçet's syndrome.

The EMBO journal·2023

Related Experiment Video

Updated: May 23, 2026

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
10:21

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification

Published on: September 21, 2011

Two-dimensional protein separation by the HPLC system with a monolithic column.

Hironobu Morisaka1, Aya Kirino, Kengo Kobayashi

  • 1Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan.

Bioscience, Biotechnology, and Biochemistry
|March 28, 2012
PubMed
Summary

A novel two-dimensional high-performance liquid chromatography system offers improved protein separation. This advanced method enhances proteomic analysis for biological research.

More Related Videos

Affinity Purification of a 6X-His-Tagged Protein using a Fast Protein Liquid Chromatography System
07:19

Affinity Purification of a 6X-His-Tagged Protein using a Fast Protein Liquid Chromatography System

Published on: April 26, 2024

Related Experiment Videos

Last Updated: May 23, 2026

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
10:21

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification

Published on: September 21, 2011

Affinity Purification of a 6X-His-Tagged Protein using a Fast Protein Liquid Chromatography System
07:19

Affinity Purification of a 6X-His-Tagged Protein using a Fast Protein Liquid Chromatography System

Published on: April 26, 2024

Area of Science:

  • Biochemistry and Analytical Chemistry
  • Proteomics and Cell Biology

Background:

  • Conventional protein separation techniques face limitations in resolution and efficiency.
  • Advancements in chromatography are crucial for comprehensive proteomic analysis.

Purpose of the Study:

  • To develop and evaluate a novel two-dimensional high-performance liquid chromatography (2D-HPLC) system for enhanced protein separation.
  • To assess the system's efficacy in proteomic analysis of yeast cell surface proteins.

Main Methods:

  • Development of a 2D-HPLC system utilizing an ion-exchange column in the first dimension and a reversed-phase monolithic column in the second dimension.
  • Application of the developed 2D-HPLC system for the separation of proteins extracted from yeast cell surfaces.
  • Evaluation of the separation efficiency compared to conventional chromatographic methods.

Main Results:

  • The developed 2D-HPLC system achieved efficient protein separation, outperforming conventional systems.
  • Successful application of the system for the proteomic analysis of yeast cell surface proteins.
  • Demonstrated high resolution and effectiveness in complex protein mixture analysis.

Conclusions:

  • The novel 2D-HPLC system provides a powerful tool for high-resolution protein separation.
  • This method significantly advances proteomic analysis capabilities, particularly for cell surface proteins.
  • The system offers a promising alternative for researchers in biochemistry and proteomics.