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Related Concept Videos

Optimizing Chromatographic Separations01:15

Optimizing Chromatographic Separations

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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...
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Principles Of Column Chromatography01:13

Principles Of Column Chromatography

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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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Types Of Column Chromatography01:29

Types Of Column Chromatography

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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.
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Ion-Exchange Chromatography01:09

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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...
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Affinity Chromatography

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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...
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Chromatography: Introduction01:10

Chromatography: Introduction

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Chromatography is a technique used to separate compounds based on differences of partitioning between two phases, the stationary phase and the mobile phase.
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Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
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Algorithms to optimize multi-column chromatographic separations of proteins.

Santiago Codesido1, Davy Guillarme1, Szabolcs Fekete1

  • 1Institute of Pharmaceutical Sciences of Western Switzerland (ISPSO), University of Geneva, CMU-Rue Michel Servet 1, 1211, Geneva 4, Switzerland; School of Pharmaceutical Sciences, University of Geneva, CMU-Rue Michel Servet 1, 1211, Geneva 4, Switzerland.

Journal of Chromatography. A
|January 10, 2021
PubMed
Summary

This study introduces an automated algorithm and downloadable software for optimizing multi-column protein separation systems. The solution enables precise control over column segments for tailored protein fractionation and analysis.

Keywords:
Column couplingMonoclonal antibodyMulti-isocratic elutionOn-column fractioningOptimizationProtein analysis

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Area of Science:

  • Biochemistry
  • Chemical Engineering
  • Computational Biology

Background:

  • Protein separation and fractionation are critical in biopharmaceutical development.
  • Optimizing multi-column chromatography systems is complex and time-consuming.
  • Existing methods lack automated solutions for complex separation challenges.

Purpose of the Study:

  • To develop a technical solution for automated optimization of multi-column systems for protein separation.
  • To provide a downloadable algorithm and software for researchers.
  • To enable flexible control over column segment length and order for tailored separations.

Main Methods:

  • Development of a novel optimization algorithm considering column segment length and order.
  • Implementation of the algorithm into user-downloadable software.
  • Testing the algorithm with representative examples for protein fractionation.

Main Results:

  • The algorithm successfully optimizes multi-column systems for various separation goals.
  • Demonstrated uniform peak distribution for improved protein analysis.
  • Achieved on-column fractionation of intact monoclonal antibodies (mAbs).
  • Showcased uniform selectivity between monoclonal antibody (mAb) sub-units.

Conclusions:

  • The developed algorithm and software offer an automated and efficient solution for multi-column protein separation.
  • This tool facilitates advanced protein fractionation and analysis, including challenging mAb separations.
  • The software provides researchers with a powerful method to optimize chromatographic processes for diverse applications.