Journal
/
Biology
/
Protein Rensing-fri metode for Binding Affinity Fastsettelse av mikroskala Thermophoresis
JoVE Journal
Biology
A subscription to JoVE is required to view this content.  Sign in or start your free trial.
JoVE Journal Biology
Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis

Protein Rensing-fri metode for Binding Affinity Fastsettelse av mikroskala Thermophoresis

DOI:
10.3791/50541-v

•

10:22 min

•

August 15, 2013

•

, , , , , ,
1Cancer and Inflammation Program,National Cancer Institute, 2Basic Science Program,SAIC-Frederick, Inc., 3Departments of Oncology and Radiation Medicine,Georgetown University Medical Center, 4Structural Biophysics Laboratory,National Cancer Institute

Chapters

  • 00:05Title
  • 01:43Preparation of Cell Lysate
  • 03:20Determination of Optimal Lysate Dilution
  • 04:02Microscale Thermophoresis Binding Studies
  • 07:05Microscale Thermophoresis Data Analysis
  • 08:25Results: Measuring the Affinity of Non-phosphorylated STAT3 Protein Binding to Oligonucleotides
  • 09:40Conclusion

Summary

Automatic Translation

Automatic Translation

Mikroskala Thermophoresis (MST) kan bli mye brukt til fastsettelse av bindende affinitet uten rensing av målet protein fra cellelysater. Protokollen omfatter overekspresjon av GFP-sammensmeltet protein, cellelyse i ikke-denaturerende betingelser, og påvisning av MST-signal i nærvær av varierende konsentrasjoner av ligand.

Tags

Protein PurificationMicroscale ThermophoresisBinding AffinityKD DeterminationGFP-fused ProteinSTAT3Cell LysateNon-denaturing ConditionsDrug DiscoveryProtein Interactions

Article

Embed

ADD TO PLAYLIST

Usage Statistics

Related Videos

Microvolume Protein Concentration Determination using the NanoDrop 2000c Spectrophotometer

The Importance of Correct Protein Concentration for Kinetics and Affinity Determination in Structure-function Analysis

Isolation of Labile Multi-protein Complexes by in vivo Controlled Cellular Cross-Linking and Immuno-magnetic Affinity Chromatography

PAR-CliP - A Method to Identify Transcriptome-wide the Binding Sites of RNA Binding Proteins

Lipid Vesicle-mediated Affinity Chromatography using Magnetic Activated Cell Sorting (LIMACS): a Novel Method to Analyze Protein-lipid Interaction

Ortogonale Protein Purification Tilrettelagt av en Small Bispecific Affinity Tag

Identifisering av Protein Samspill Partners Bruke Tandem Affinity Rensing

Real-time analyser av Retinol Transport av membranen Receptor av Plasma Retinol Binding Protein

A Sensitive and Specific Quantitation Method for Determination of Serum Cardiac Myosin Binding Protein-C by Electrochemiluminescence Immunoassay

Identifisere Protein-protein samhandling i<em> Drosophila</em> Voksen Heads av Tandem Affinitetsrensing (TAP)

Read Article