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Analysis Kinetics of Disease Biomarker TNF-α by BLI (CAT#: STEM-MB-0246-CJ)

Introduction

In recent years, biomarkers have been extensively investigated for use in disease diagnosis, treatment monitoring, and assessing the prognosis of diseases. Once released from the source of the disease, biomarkers, such as proteins, nucleic acids, and metabolites, may spread to the blood, urine, tears, and saliva. In particular, protein biomarkers in human blood, such as tumor necrosis factor-α (TNF-α), have been widely used as diagnostic indicators for cancer, Alzheimer’s disease, rheumatoid arthritis, and diabetic eye disease. The concentrations of TNF-α in blood are generally lower than 40 pg/mL for healthy humans, while significantly increase to about 300 pg/mL for patients with severe diseases.




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

Oncology & Cancer; Immunology/Inflammation; 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: Gator® Bio-Layer Interferometry (BLI)
• Sample Type: DNA, RNA, Protein, Antibodies, Peptides, Small Molecules
• Optionals: Recombinant Human Tumor Necrosis Factor-α (TNF-α), Recombinant Human Connective tissue Growth Factor (CTGF), Human Serum Albumin (HSA), Thrombin, Bovine Serum Albumin (BSA), Vascular Endothelial Growth Factor165 (VEGF165)
lipocalin 1 (LCN 1), Angiopoietin-like 4 (ANGPTL4)