Desthiobiotin-streptavidin Affinity
Desthiobiotin-streptavidin Affinity - Related Videos
11:11
Desthiobiotin-Streptavidin-Affinity Mediated Purification of RNA-Interacting Proteins in Mesothelioma Cells
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Cited by 6 •
2018
Desthiobiotin labeling of a synthetic 25-nucleotide RNA oligo, which contains an adenine-rich element (ARE) motif, allows specific binding of cytosolic ARE-binding protein.
05:07
RNA-Protein Pull-Down Assay to Isolate RNA-Binding Proteins via Affinity Extraction
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2025
This video demonstrates an in vitro RNA pull-down assay to identify RNA-binding proteins (RBPs), which interact with the adenylate-uridylate-rich element (ARE) sequences in mRNA. The target RBPs from a cell lysate are mixed with an RNA probe to form RNA-protein complexes. The complexes are isolated via affinity purification utilizing the affinity of the desthiobiotin label of the RNA probe to a streptavidin-labeled magnetic bead.
04:51
Streptavidin-Affinity Grid Fabrication for Cryo-Electron Microscopy Sample Preparation
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Cited by 15 •
2023
A step-by-step protocol for fabricating streptavidin affinity grids is provided for use in structural studies of challenging macromolecular samples by cryo-electron microscopy.
03:41
Analyzing Influenza Virus Internalization Using a Streptavidin Blocking Assay
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2026
Source: Pohl, M. O. & Stertz, S. Measuring Attachment and Internalization of Influenza A Virus in A549 Cells by Flow Cytometry. J. Vis. Exp. (2015)This video demonstrates the use of a streptavidin blocking assay to distinguish surface-bound from internalized influenza virus in human lung epithelial cells. Comparing fluorescence signals shows that a higher intensity after incubation indicates successful viral internalization.
06:34
Tandem Affinity Purification Assay to Study Protein-Protein Interactions
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2025
This video demonstrates tandem affinity purification — a technique to purify protein complexes from eukaryotic cells to study protein-protein interaction. The complex contains two proteins labeled with different epitope tags. Upon the purification of the complex using two resins with an affinity for the two different tags in a sequential manner, the presence of both proteins in the elute confirms the interaction between the two.
FAQs
Desthiobiotin occupies a biotin-binding pocket on streptavidin through a specific noncovalent interaction. Because this interaction is weaker than permanent biotin-streptavidin binding, free biotin can compete for the same pocket and displace the captured desthiobiotin-labeled material. This competitive release provides a way to recover biomolecules without relying on harsh conditions that may disrupt their structure or activity.
Lower affinity makes the capture interaction reversible rather than effectively permanent. Streptavidin can still retain labeled proteins, nucleic acids, or molecular complexes during the capture step, while free biotin can later promote release. This balance is useful when the recovered material must remain suitable for downstream biochemical or analytical studies.
Both systems use the specific binding pockets of streptavidin, but their practical behavior differs during recovery. Biotin-streptavidin capture is described as permanent, whereas desthiobiotin permits competitive displacement by free biotin under mild elution conditions. The reversible format therefore offers greater control over release and can reduce the need for denaturing treatment after capture.
Free biotin acts as the competing ligand during elution. It binds to streptavidin’s available biotin-binding pockets and displaces desthiobiotin from the interaction, allowing the associated biomolecule or complex to be recovered. This mechanism is central to mild release because it changes the binding competition rather than requiring harsh disruption of the captured sample.
A typical workflow first brings a desthiobiotin-associated target into contact with streptavidin so the labeled protein, nucleic acid, or molecular complex is captured. After the desired material has been isolated, free biotin is introduced to compete for streptavidin binding sites and release the target. The recovered sample can then proceed to biochemical or analytical analysis.
This approach is especially useful when a pull-down assay or purification requires both selective capture and subsequent recovery of the retained material. It can be applied to labeled proteins, nucleic acids, and molecular complexes, including samples whose structure or activity may be affected by harsh elution. Reversible release supports examination of the isolated material in downstream studies.
The system can support isolation of desthiobiotin-associated proteins, nucleic acids, and molecular complexes. Its value extends beyond simply retaining a target: the material can also be released through competition with free biotin under mild conditions. This combination makes the method relevant to affinity purification, pull-down experiments, and studies that analyze recovered biomolecular interactions.