Fabrication Of Lipid Bilayer Arrays And The Migration Of Charged Lipids Within Lipid Bilayer In An Electric Field

The supported lipid bilayer arrays prepared based on bionic principle could be used to study the structures and functions of cell membrane,which has become a rising star in biofilm research area.The control of charged species in the supported lipid bilayer using extra electric field could provide theoretical and practical guidance for studies of structures and functions of charged species,such as membrane proteins.However,in the present studies,there are still many questions to be solved,including the relationship between the supported lipid bilayer formation and surface wetting property,the height control of the barriers of lipid bilayer arrays at the nanoscale,the control of charged species migration within the patterned supported lipid bilayer in an electric field.To solve these problems,following studies were carried out.A surface with chemical gradient was created using space limited plasma oxidization method,which was used to study the relationship between the supported lipid bilayer formation and surface wetting properties.Nanoscale controlled barrier was built with layer-by-layer of polyelectrolyte in combination with ultraviolet lithography method,which was used to prepare supported lipid bilayer arrays.Theoretical calculations and experiments were done to study the directed migration of charged lipids within the special patterned supported lipid bilayer arrays in an AC electric field.The main contents of this thesis are described as follows.The surface contact angle gradient from about 108 ° to about 5 ° was created using space limited plasma oxidization method.The giant unilamellar vesicles(GUVs)were used to study the relationship between the supported lipid bilayer formation and surface wetting property.The results of fluorescence microscopy and atomic force microscopy indicated that lipid monolayer was formed on the surface with contact angle from about 61 ° to about 108 °;the monolayer patches were formed on the surface with contact angle between about 55 ° and about 60 °;GUVs were intactly attached on the surface with contact angle from about 28 ° to about 54 °;lipid bilayer patches were formed on the hydrophilic area with the contact angle to be about 5 °.In addition,small unilamellar vesicles(with SUVs)could form a ho mogeneous supported lipid bilayer on the surface with contact angle about 5 °.It can be concluded that supported lipid bilayer should be formed on the substrate with contact angle about 5°,and in order to obtain a continuous support lipid bilayer SUVs sh ould be used for the vesicle fusion method.Subsequently,fluorescence recovery after photobleaching technique(FRAP),cyclic voltammetry and impedance spectroscopy were used to study the influence of fluidity and permeability of lipid bilayer by baicalin and baicalein.The results showed that baicalin had little effect on the properties of lipid bilayer while the baicalein had a more significant impact.When the concentration of baicalein is 25 μmol/L,the diffusion coefficient of charged lipids in lipid bilayer decreased from 1.93 ± 0.02 μm2/s to 1.34 ± 0.02 μm2/s.Furthermore,the permeability of lipid bilayer increased as the rising of baicalein concentration.The responses of K3Fe(CN)6 on lipid bilayer membrane modified Pt electrode linearly increased.Based on the study of supported lipid membrane preparation,the supported lipid bilayer arrays were built.First the polyelectrolyte film formed by layer-by-layer method was treated with ultraviolet lithography method to create the patterned substrate where the polyelectrolyte acted as barriers.FRAP results indicated the supported lipid bilayer prepared by this method had very good mobility.Meanwhile,the negative charged NBD PE within the supported lipid bilayer could move at 10 V/cm DC electric field.Based on the Browine ratchet mechanisms,an asymmetric square wave AC electric field and rotating electric field were used to control the charged lipids movement within the supported lipid bilayer.From COMSOL finite element analysis and experimental results,it could be found that there were some key factors effecting the charged lipid movement under the electric field: asymmetry of ratchets,ratchet height vs.time of reversed electric field,phase angle of rotating AC electric field and period.The results showed that the bigger asymmetry factor caused higher the efficiency of the movement of charged lipids within the supported lipid bilayer;the charged lipids were enriched in the last rachet under 62 V/cm electric field with 16 min reversed electric field;the parameters h’/h was used to represent the influence of the time of reversed electric field and ratchet height.The lateral concentrate of charged lipids requires appropriate h’/h value,i.e.about 1;the charged lipids could be concentrated most effectively when the phase difference of rotating AC electric field is π/2 and the period is 9600 s.In this thesis,surface chemical gradient substrates prepared with space limited plasma oxidization method were used to clarify the relationship between the s urface wetting property and the formation of support lipid membranes.The combination of layer-by-layer of the polyelectrolyte multilayers with ultraviolet lithography technology could break through the traditional method to build a new method to prepare supported lipid bilayer with controlled barrier in nano scale.Both a finite element analysis model and an experimental setup have been used to investigate the role of key parameters for the operation of these ratchets.This thesis effectively solved the problems of charged species migration in the support lipid bilayer arrays,and provided a theoretical basis for the control of the motion of charged species within supported lipid bilayer,which has potentials in study of the structure and function of membrane proteins.