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Related Concept Videos

Ion Exchange01:17

Ion Exchange

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Ion exchange chromatography separates charged molecules from a solution by reversibly exchanging them with mobile, or 'active', ions associated with the oppositely charged stationary phase. This method can be used to separate ions, soften and deionize water, and purify solutions. The polymers comprising the ion-exchange column are high-molecular-weight and chemically stable polymers, crosslinked to be porous and essentially insoluble. They are also functionalized with either acidic or...
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Principles Of Column Chromatography01:13

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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

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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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High-Performance Liquid Chromatography: Elution Process01:05

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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...
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Capillary Electrophoresis: Applications01:30

Capillary Electrophoresis: Applications

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Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
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Titration of Polyprotic Base with a Strong Acid01:18

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The titration of a polyprotic base such as sodium carbonate with a strong acid such as hydrochloric acid results in two equivalence points on the titration curve. At the first equivalence point, the carbonate ions in the base are completely converted to bicarbonate ions. The second equivalence point corresponds to the complete conversion of bicarbonate ions to carbonic acid, which dissociates into carbon dioxide and water. The region before the first equivalence point corresponds to the...
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Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
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Obtaining acidic and basic charge variants using a twin-column continuous chromatography system.

Gaoya Yuan1, Xudong Zhang1, Yuanyi Zhang1

  • 1Technology and Process Development (TPD), WuXi Biologics, 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai, 200131, China.

Protein Expression and Purification
|December 18, 2022
PubMed
Summary
This summary is machine-generated.

Producing high-purity acidic and basic charge variants of monoclonal antibodies (mAbs) is crucial for characterization. A novel continuous chromatography method efficiently separates and collects these critical mAb variants, enabling detailed analysis.

Keywords:
Cation exchange (CEX) chromatographyCharge variantsContinuous chromatographyMonoclonal antibody (mAb)Twin-column

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Area of Science:

  • Biopharmaceutical Process Development
  • Protein Chemistry
  • Analytical Chemistry

Background:

  • Recombinantly produced monoclonal antibodies (mAbs) exhibit charge heterogeneity, with acidic and basic variants being common.
  • Accurate characterization of these mAb charge variants necessitates obtaining sufficient quantities of highly pure acidic and basic species.
  • Existing methods may not efficiently provide the required amounts of purified variants for comprehensive analysis.

Purpose of the Study:

  • To develop an efficient approach for the continuous separation and collection of acidic and basic charge variants of monoclonal antibodies.
  • To achieve high purity (>90%) and sufficient quantities of separated mAb charge variants.
  • To facilitate the detailed characterization of monoclonal antibody charge variants.

Main Methods:

  • Optimization of batch-mode cation exchange (CEX) chromatography to determine optimal load density and linear salt gradient elution conditions.
  • Development of a stepwise elution protocol derived from the optimized linear gradient elution.
  • Implementation of a customized twin-column continuous chromatography system utilizing the stepwise elution protocol for persistent variant production.

Main Results:

  • Established optimal conditions for separating acidic and basic charge variants using batch CEX chromatography.
  • Successfully developed and implemented a stepwise elution protocol for continuous separation.
  • Achieved efficient and continuous production of acidic and basic mAb charge variants with high purity (>90%) using a twin-column system.

Conclusions:

  • The developed continuous chromatography approach effectively generates sufficient amounts of high-purity mAb charge variants.
  • This method significantly aids in the essential characterization of acidic and basic mAb charge variants.
  • The twin-column continuous system offers a robust solution for producing purified mAb variants for analytical purposes.