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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...
Optimizing Chromatographic Separations01:15

Optimizing Chromatographic Separations

Optimizing chromatographic separations is crucial for obtaining clean separations in a minimum amount of time. Optimization is required for several factors, including kinetic effects related to band broadening, plate height, capacity factor, and separation factor.
Band broadening refers to spreading solute bands as they travel through the column. This broadening can impact resolution. Plate height (H) represents the length required for one theoretical plate. A lower plate height corresponds to...
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...
Silica Gel Column Chromatography: Overview01:10

Silica Gel Column Chromatography: Overview

Silica gel column chromatography is a technique for separating compounds using a column packed with silica gel as the stationary phase. This method relies on differences in the polarity of compounds. Based on their polarities, compounds move between the stationary phase (silica gel) and the mobile phase (the solvent), forming discrete bands in the column.
Polar components tend to bind strongly to the silica gel, causing them to move slowly through the column. In contrast, nonpolar compounds...
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...
Chromatographic Resolution01:15

Chromatographic Resolution

In chromatography, a solute moves through a chromatographic column and tends to spread, forming a Gaussian-shaped band. The longer the solute spends in the column, the broader the band becomes. The broadening can lead to overlaps within the column, affecting separation effectiveness.
The effectiveness of separation can be evaluated by determining the level of separation between two neighboring peaks in a chromatogram, which represents the individual components of a sample.
In chromatography,...

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Related Experiment Video

Updated: May 25, 2026

Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study
07:53

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

Published on: August 16, 2019

Improving affinity chromatography resin efficiency using semi-continuous chromatography.

Ekta Mahajan1, Anupa George, Bradley Wolk

  • 1Technical Development Engineering, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA. ektam@gene.com

Journal of Chromatography. A
|January 24, 2012
PubMed
Summary

Continuous chromatography using three Protein A columns significantly reduces resin and buffer use in monoclonal antibody purification. This periodic counter-current chromatography (3C PCC) method offers substantial cost savings compared to traditional batch processing.

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Last Updated: May 25, 2026

Activated Cross-linked Agarose for the Rapid Development of Affinity Chromatography Resins - Antibody Capture as a Case Study
07:53

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

Published on: August 16, 2019

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High-Throughput Expression and Purification of Human Solute Carriers for Structural and Biochemical Studies

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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

Area of Science:

  • Biotechnology
  • Chemical Engineering
  • Biopharmaceutical Manufacturing

Background:

  • Protein A affinity chromatography is a standard method for monoclonal antibody (MAb) purification from harvested cell culture fluid (HCCF).
  • High costs associated with Protein A resin represent a significant portion of raw material expenses in MAb manufacturing.
  • Current manufacturing often employs single-column batch processing, leading to potential inefficiencies in resin utilization.

Purpose of the Study:

  • To evaluate the efficacy of a three-column periodic counter-current chromatography (3C PCC) system for continuous MAb purification.
  • To assess the potential for cost reduction through decreased resin volume and buffer consumption.
  • To compare the yield and quality of MAbs purified using the continuous 3C PCC method versus traditional batch processing.

Main Methods:

  • Experiments were conducted using three 1-ml Hi-Trap™ MabSelect SuRe™ columns on a modified ÄKTA™ system.
  • The 3C PCC principle was applied, with columns loaded sequentially until a 70% breakthrough point was reached.
  • Unbound protein from one column was loaded onto the next, ensuring continuous capture of MAbs.
  • Product yield and quality were assessed and compared to a batch purification process.

Main Results:

  • Continuous operation using 3C PCC demonstrated a potential reduction in resin volume and buffer consumption by approximately 40%.
  • The continuous process led to a reduction in overall processing time compared to batch methods.
  • Alternative methods like effluent recycling and increased residence time offered similar cost benefits but required longer processing times.

Conclusions:

  • The three-column periodic counter-current chromatography (3C PCC) process offers a viable strategy for reducing costs in MAb purification.
  • Continuous processing via 3C PCC can significantly decrease resin and buffer requirements, leading to economic advantages.
  • While more complex, the 3C PCC system presents a compelling alternative to batch processing for large-scale MAb manufacturing.