Immobilization of Lipophilic and Amphiphilic Biomarker on Hydrophobic Microbeads

Background: Lipids and amphiphilic molecules are ubiquitous and play a central role in cell signalling, cell membrane structure, and lipid transport in the human body. However, they also appear in many diseases such as atherosclerosis, cardiovascular diseases, infections, inflammatory diseases, cancer, and autoimmune diseases. Thus, it is necessary to have detection systems for lipids and amphiphilic molecules. Microbeads can be one of these systems for the simultaneous detection of different lipophilic biomarkers. Methods: Based on the fundamentals of microbead development, novel hydrophobic microbeads were produced. These not only have a hydrophobic surface, but are also fluorescently encoded and organic solvent resistant. The challenge after the development of the hydrophobic microbeads was to immobilise the amphiphilic molecules, in this study phospholipids, on the microbead surface in an oriented direction. After successful immobilisation of the biomarkers, a suitable antibody based detection assay was established. Results: By passive adsorption, the phospholipids cardiolipin, phosphatidylethanolamine and phosphatidylcholine could be bound to the microbead surface. With the application of the enzymes phospholipase A2 and phospholipase C, the directional binding of the phospholipids to the microbead surface was demonstrated. The detection of directional binding indicated the need for the hydrophobic surface. Microbeads with no hydrophobic surface bound the phospholipids non-directionally (with the hydrophilic head) and were thus no longer reactively accessible for detection. Conclusion: With the newly developed hydrophobic, dual coded and solvent stable microbeads it is possible to bind amphiphilic biomolecules directionally onto the microbead surfaces.