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Analysis Biomolecular Interactions of SARS-CoV-2 N Protein with Antibody by BLI (CAT#: STEM-MB-0154-CJ)

Introduction

Besides being the capsid protein of the virus, the N-protein of many coronaviruses is known to double up as a regulatory protein. The N-protein of the SARS-CoV has been shown to modulate the host cellular machinery in vitro, thereby indicating its possible regulatory role during its viral life cycle. SARS-CoV-2 N protein can promote NLRP3 inflammasome activation to induce hyperinflammation. Arginine methylation of SARS-Cov-2 nucleocapsid protein regulates RNA binding, its ability to suppress stress granule formation, and viral replication.




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

Immunology/Inflammation; Toxicology; Virology

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: PCR Amplifier, polyacrylamide Gel Electrophoresis (PAGE)