Affinity-based Separation
Affinity-based Separation - Related Videos
03:07
Electron Affinity
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2020
The electron affinity (EA) is the energy change for adding an electron to a gaseous atom to form an anion (negative ion). This process can be either endothermic or exothermic, depending on the element. Many of these elements have negative values of EA, which means that energy is released when the gaseous atom accepts an electron. However, for some elements, energy is required for the atom to become negatively charged, and the value of their EA is positive. Just as with ionization energy,...
01:41
Affinity and Avidity
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2019
Overview Antibodies bind to toxins or substances on the surface of cells, bacteria, viruses, or fungi. The substance is called an antigen, and the precise binding site is the epitope. The strength of the antibody-epitope interaction is called affinity. When an antibody binds an antigen by multiple epitopes, the cumulative strength of the interaction is called avidity. The strength of the interaction influences the elicited immune response. The Adaptive Immune System Increases Efficiency by...
06:08
Microfluidic Acoustophoresis for Flowthrough Separation of Gram-Negative Bacteria using Aptamer Affinity Beads
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2022
This paper describesthe fabrication and operation of microfluidic acoustophoretic chips using the microfluidic acoustophoresis technique and aptamer-modified microbeads that can be used for fast, efficient isolation of Gram-negative bacteria from a medium.
02:21
Purification of Viral Integrase Using Heparin Affinity Chromatography
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2026
Source: Lopez Jr., M. et al. Detection and Removal of Nuclease Contamination During Purification of Recombinant Prototype Foamy Virus Integrase. J. Vis. Exp. (2017)This video demonstrates the procedure for purifying viral integrase from bacterial nuclease contamination using heparin Sepharose column chromatography followed by SDS-PAGE analysis to confirm protein purity.
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.
FAQs
Selectivity comes from matching an immobilized ligand to a binding feature on the intended target. Because the mixture contains components that do not recognize that ligand, they can be removed during washing while the bound material remains associated with the support. The quality of this molecular match largely determines which species are retained and how clean the recovered fraction becomes.
Release conditions determine whether the target leaves the support without losing the usefulness of the separation. Adjusting pH or salt concentration can weaken the binding interaction, while a competing ligand can displace the captured material. These alternatives make it possible to recover the target after unwanted components have been washed away, producing a separate fraction for further analysis.
Its value in complex biological samples comes from combining molecular recognition with physical retention. The ligand retains the species that matches it on the solid support, whereas other mixture components are removed before recovery. This can raise sample purity and reduce the number of processing steps needed before biochemical analysis or characterization.
Researchers first attach a target-recognizing ligand to a solid support and expose that support to the biological mixture. After binding, they wash away unbound material, then alter pH, salt concentration, or add a competing ligand to release the retained target. The recovered fraction can then be collected for subsequent biological or biochemical work.
The approach is useful when a biological sample contains a target that must be isolated selectively, including proteins, antibodies, nucleic acids, or cells. Depending on the captured material, the resulting fraction can support biochemical analysis, biomarker studies, therapeutic production, or molecular characterization. Its main practical benefit is obtaining a more suitable sample with fewer processing steps.
In biology, the method provides information beyond sample cleanup. Which material binds to the immobilized ligand, remains after washing, and is released during recovery can help researchers examine recognition between molecular partners. This makes the technique relevant to characterizing molecular interactions as well as preparing purified material for downstream biological studies.