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Related Concept Videos

Affinity Chromatography01:03

Affinity Chromatography

Affinity chromatography is a powerful technique extensively utilized for separating and purifying specific biomolecules from complex mixtures. It capitalizes on the highly selective binding between an analyte and its counterpart, such as antibody-antigen interactions. The counterpart is immobilized on the stationary phase, forming an affinity column. The stationary phase typically consists of solid support, such as agarose or porous glass beads, immobilizing the affinity ligand. The mobile...
Ion-Exchange Chromatography01:09

Ion-Exchange Chromatography

Ion-exchange chromatography, or IEC, is a technique for separating ions based on their affinity for the stationary phase. The stationary phase is a cross-linked polymer resin with covalently attached ionic functional groups. The functional groups can be either positively charged (cation exchangers) or negatively charged (anion exchangers). A cation exchanger consists of a polymeric anion and active cations, while an anion exchanger is a polymeric cation with active anions. The choice of...
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...
Chromatographic Methods: Classification01:12

Chromatographic Methods: Classification

Chromatographic techniques are classified in three ways: the classification is based on the physical state of the stationary and mobile phases, how the mobile phase and the stationary phase contact each other, or through the chemical or physical processes that isolate the components of the sample. Typically, the mobile phase is either a liquid or gas, while the stationary phase is either a solid or a liquid layer applied to a solid surface.
Chromatographic techniques are typically named by...
Chromatography: Introduction01:10

Chromatography: Introduction

Chromatography is a technique used to separate compounds based on differences of partitioning between two phases, the stationary phase and the mobile phase.
The phase in which the compounds linger or on which the compounds adsorb is called the stationary phase, whereas the mobile phase is the solvent that carries the solutes to be analyzed. In traditional column chromatography, the mixture flows through the stationary phase, and the compounds partition between the stationary and mobile phases...
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...

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Related Experiment Video

Updated: Jun 19, 2026

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

Affinity chromatography: general methods.

Marjeta Urh1, Dan Simpson, Kate Zhao

  • 1Promega Corporation, Madison, Wisconsin, USA.

Methods in Enzymology
|November 7, 2009
PubMed
Summary
This summary is machine-generated.

Affinity chromatography purifies molecules using specific biological interactions. Optimizing ligand selection and attachment chemistry is crucial for efficient purification and high recovery rates.

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Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study
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Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study

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Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow
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Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow

Published on: January 8, 2014

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Last Updated: Jun 19, 2026

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

Published on: April 26, 2024

Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study
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Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study

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Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow
10:33

Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow

Published on: January 8, 2014

Area of Science:

  • Biochemistry
  • Chromatography
  • Molecular Biology

Background:

  • Affinity chromatography is a powerful technique for purifying specific molecules from complex mixtures.
  • It relies on highly specific, reversible biological interactions between molecules, like enzyme-substrate or antibody-antigen.
  • These interactions are leveraged by immobilizing one molecule (ligand) onto a matrix to capture the target molecule.

Purpose of the Study:

  • To discuss key factors in selecting affinity ligands and matrices for effective purification.
  • To highlight the importance of understanding molecular interactions for successful affinity purification.
  • To describe recent advancements in affinity chromatographic materials.

Main Methods:

  • Utilizing specific biological interactions (e.g., enzyme-substrate, antibody-antigen) for molecular purification.
  • Immobilizing affinity ligands onto solid matrices to create stationary phases.
  • Developing and selecting appropriate matrices and attachment chemistries for ligand coupling.

Main Results:

  • Affinity purification offers significant time savings and high purification factors (hundreds-fold or more).
  • Commercial availability of common ligands simplifies the process.
  • Newer matrices offer improved features over traditional materials.

Conclusions:

  • Successful affinity purification hinges on careful selection of ligands and matrices, and understanding binding interactions.
  • Optimization of purification protocols is essential for efficient target molecule capture and recovery.
  • Advancements in materials enhance the capabilities of affinity chromatography.