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Analysis Biomolecular Interactions of GST-HP1γ Proteins with Immobilised H3 by BLI (CAT#: STEM-MB-0227-CJ)

Introduction

HP1 was originally described as a dominant suppressor of position-effect variegation in Drosophila melanogaster. The mammalian HP1 protein family consists of three members: HP1α, β and γ. As the main reader of repressive histone marks H3K9me2/3, HP1 is a key factor in heterochromatin formation and maintenance. And HP1γ is predominantly found in euchromatin, within the transcribed regions of active genes. HP1γ plays important roles in developmental processes and cell fate decisions. For instance, depletion of HP1γ in mESCs results in endodermal and neuronal differentiation defects. HP1γ can regulat transcription during differentiation.




Principle

Bio-Layer Interferometry (BLI) is an optical technique for measuring macromolecular interactions by analyzing interference patterns of white light reflected from the surface of a biosensor tip. BLI experiments are used to determine the kinetics and affinity of molecular interactions. In a BLI experiment, one molecule is immobilized to a Dip and Read Biosensor and binding to a second molecule is measured. A change in the number of molecules bound to the end of the biosensor tip causes a shift in the interference pattern that is measured in real-time.

Applications

Cardiology

Procedure

1. Detect Buffers and prepare samples. BLI experiments are set up with one molecule immobilised on the surface of the biosensor (load sample) and a second molecule in solution (the analytical sample).
2. Fix the load sample on the biocompatible biosensor while the analytical sample is in solution.
3. The biosensor tip is immersed in the solution so that the target molecule begins to bind to the analysis sample.
4. Set up and run the BLI experiment. Molecules bound to or dissociated from the biosensor can generate response curves on the BLI system; unbound molecules, changes in the refractive index of the surrounding medium or changes in flow rate do not affect the interferogram pattern.
5. Collect and analyse data on the BLI's system.

Materials

• Equipment: Fortebio Bio-Layer Interferometry (BLI)
• Sample Type: DNA, RNA, Protein, Antibodies, Peptides, Small Molecules
• Optionals: The vector pSB7