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Analysis Biomolecular Interactions of Pfs25 Monoclonal Antibodies (mAbs) to Pfs28 by BLI (CAT#: STEM-MB-0166-CJ)

Introduction

Pfs25 is a leading TBV candidate expressed during the sexual stages of the parasite lifecycle and plays a role in ookinete survival in the mosquito midgut as well as traversal through the midgut epithelium. Pfs25 has progressed through extensive clinical investigation to assess its safety and efficacy. Several approaches have been explored to increase the immunogenicity of Pfs25, including virus-like particle (VLP) display, nanoparticle display, conjugation to Pseudomonas aeruginosa Exoprotein A (EPA) and formulation in diverse adjuvants.<br /><br />Pfs28 is a glycosylphosphatidylinositol-anchored protein expressed on the surface of zygotes and ookinetes and plays a similar role as Pfs25 in ookinete survival and traversal through the midgut. This function of Pfs28 may be mediated through its interactions with basal lamina proteins surrounding the mosquito midgut. Pfs28 and Pfs25 are thought to play partially redundant roles in parasite survival within the mosquito.




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; 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: Fortebio Bio-Layer Interferometry (BLI)
• Sample Type: DNA, RNA, Protein, Antibodies, Peptides, Small Molecules
• Optionals: Pfs25 (PF3D7_1031000), Pfs28