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An outlook with a discussion of the current limitations to design screening devices for membrane proteins is in Chapter 7.Future developments of the work presented in this thesis include: addressing the switch on/off of individual pores for controlled transport across selected pores; analyzing the transport of ions and low-molecular-weight drug candidates per single nanopore; and test the universal applicability of the designed assay platforms by integrating different types of membrane proteins.
It was also demonstrated that the protein density could be varied in a wide range without impairing the formation of the lipid bilayer.
Although improvements are needed for the electrochemical measurements of membrane protein activity, yet the potential application of the integrated platform for ion channel protein assays is demonstrated.
Several properties of the smart PMAA brush contributed to the functionality of such protein assays.
Controlled transport of ions and molecules through nanopores, support of pore-spanning artificial lipid bilayers and positioning of membrane proteins over the pores were achieved with the explored PMAA-brush functionalized platforms.
The formation of the suspended artificial lipid bilayers was achieved by spontaneous liposome rupture and fusion.
Thesis Transport Aids Essay Conclusion
Enhanced membrane resistance as determined by electrochemical impedance spectroscopy (EIS) and free diffusion of dyed lipids observed as fluorescence recovery after photobleaching (FRAP) confirmed the presence of the lipid bilayer.All the results presented in this chapter rationalize the use of nanoporous platforms functionalized with PMAA brushes for controlled gating applications, for example as electrochemical (bio)sensors or nanoscale valves.In Chapter 6, a description is provided discussing functionalization of silicon-based nanopore array chips functionalized with PMAA brush, which were used as supports for pore-spanning lipid bilayers.The shift is due to ion confinement effects in the dense brush, and brings the conformational switching of PMAA brushes closer to physiological conditions.This is relevant for biologically oriented applications in miniaturized devices with rapid response times.The functionalized pores had higher electrical resistance when the brush was in a neutral state and lower resistance when the brush was charged.The results of these measurements, combined with diffusion experiments using a fluorescent dye to investigate mechanical gating, demonstrated the controlled transport of ions and small molecules across the pores.As additional review of the literature background for the work presented in this thesis, an overview on pore-spanning lipid bilayers for functional assays of membrane proteins is presented in Chapter 3.The role of membrane proteins as drug targets is introduced and the current limitations in the development of such assays are discussed.Students have to prepare the draft of the thesis by the fourth week of the semester.Based on this draft the Consultant composes the specification sheet of thesis, which contains the final title, 3-5 bullet points about the content, name of the External Supervisor and Final Examination subjects.