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Analysis Kinetics of CTX-MI in Tap Water by BLI (CAT#: STEM-MB-0276-CJ)

Introduction

Conus species are carnivorous gastropods living in tropical and subtropical oceans. Their venom ducts and poison glands secrete varieties of toxic polypeptides during predation and defense, which are called conotoxins (CTXs). Enormous gene families, abundant cysteines, and various post-translational modifications contribute to the diversity of CTXs. As a class of neurotoxins, CTXs act on a variety of ion channels and receptors throughout the nervous system, such as acetylcholine receptors and voltage-gated Na+ channels, with high selectivity and potency. The sting of cone snails can cause paralysis, dizziness, convulsion, respiratory failure, or even death. Alpha-conotoxin MI (CTX-MI), which was first isolated from Conus magus, is considered as the most venomous species. CTX-MI is a short peptide of 14 amino acids (GRCCHPACGKNYSC-NH2), which forms two disulfide bonds (Cys3-Cys8 and Cys1-Cys14) and selectively inhibits muscular acetylcholine receptors.




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

Neurobiology/Neurodegeneration; Immunology/Inflammation; Toxicology; Pharmacology

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: Sartorius Bio-Layer Interferometry (BLI)
• Sample Type: DNA, RNA, Protein, Antibodies, Peptides, Small Molecules
• Optionals: CTX-GI, CTX-MI, OA, STX, TTX