Chromatography Purification
Chromatography Purification - Related Videos
09:18
Purification of a Total Lipid Extract with Column Chromatography
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2023
Source: Laboratory of Jeff Salacup - University of Massachusetts Amherst The product of an organic solvent extraction, a total lipid extract (TLE), is often a complex mixture of hundreds, if not thousands, of different compounds. The researcher is often only interested in a handful of compounds. The compounds of interest may belong to one of several classes of compounds, such as alkanes, ketones, alcohols, or acids (Figure 1), and it may be useful to remove the compound classes to which it does...
10:21
Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
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Cited by 17 •
2011
An automated method for identifying suitable hydrophobic interaction chromatography (HIC) media to be used in the process of protein purification is presented. The method utilizes a medium-pressure liquid chromatography system including automated buffer blending, dynamic sample loop injection, sequential column selection, multi-wavelength analysis, and split fraction eluate collection.
02:54
An Affinity Chromatography Technique for the Purification of a Recombinant Bacterial Protein
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2025
Source: Becker, B., et al. A Simple Fluorescence-based Reporter Assay to Identify Cellular Components Required for Ricin Toxin A Chain (RTA) Trafficking in Yeast. J. Vis. Exp. (2017).This video demonstrates the purification of a polyhistidine-tagged recombinant protein using nickel-based affinity chromatography. It outlines key steps, including bacterial lysis by sonication, clarification and filtration of the lysate, and selective binding and elution of the target protein using imidazole.
07:51
Chromatography-based Biomolecule Purification Methods
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2023
In biochemistry, chromatography-based purification methods are employed to isolate compounds from a complex mixture. Two such methods used commonly by biochemists are size-exclusion chromatography and affinity chromatography. In size-exclusion chromatography, a column packed with porous beads separates components of a mixture based on size. On the other hand, affinity chromatography allows for a more specific separation of biomolecules by using a column that is composed of stationary phase,...
03:03
An Affinity Chromatography Technique for the Purification of Monoclonal Antibodies
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2025
This video demonstrates a technique for purifying monoclonal antibodies from the culture supernatant obtained from an antibody-producing cell culture. The process involves using a matrix of crosslinked agarose beads with conjugated Protein A, which immobilizes the antibodies in the supernatant on the beads, allowing for effective separation from contaminants. By applying an elution buffer with low pH, the protein A-antibody interaction is loosened, facilitating the collection of...
FAQs
Separation improves when the system creates a meaningful difference in movement between the target and contaminants. Polarity, charge, molecular size, and binding affinity determine how each component interacts with the stationary and mobile phases. Engineering teams use those differences to select a strategy that distinguishes the desired product from chemically or physically similar impurities.
Technique selection follows the property that most clearly distinguishes the target from unwanted components. Ion-exchange focuses on charge, size-exclusion on molecular size, affinity on binding affinity, and liquid chromatography can be used within purification workflows. Matching the technique to the available difference helps direct the separation toward the desired molecule.
Reproducibility improves when a purification workflow repeatedly exploits the same distinguishing property, such as charge, size, polarity, or binding affinity. A technique chosen without regard to the target and contaminants may not separate them effectively. In engineering workflows, consistent separation supports dependable product quality, improved process efficiency, and more predictable performance.
A basic workflow begins by identifying the target and the property that separates it from contaminants. The mixture is then passed through a system containing stationary and mobile phases, allowing components to move at different rates. The separated components can subsequently be isolated and assessed as part of purification or quality-control work.
Engineering applications span pharmaceutical production, biomolecule processing, chemical manufacturing, and industrial products. In these settings, the method can serve both purification and quality control, helping separate target materials from contaminants while supporting product purity. Its relevance extends from laboratory workflows to manufacturing processes where efficiency and reproducibility matter.
Key outcomes include the purity of the product, the efficiency of the process, and the reproducibility of the workflow. These measures indicate whether the selected separation approach adequately distinguishes the target from contaminants and whether it can support consistent laboratory or manufacturing operations. Quality-control use also provides a basis for evaluating pharmaceutical and industrial products.