Monitoring The Adsorption Behavior And Interaction Of Macromolecules?Nanoparticles In Microchannels With Pulsed Streaming Potential Measurement

With the development of proteomics and treatment of human diseases,investigation on the adsorption of macromolecules such as proteins,polyelectrolytes,and nanomaterials and their interactions a theoretical basis and critical to the development of physiological process involving these materials.Under non-labeled conditions with physiological buffers to monitor the interaction between macromolecules with other materials is necessary.Surface charge is the main physicochemical property of molecules,the change of surface charge will influence other properties of molecules such as the adsorption onto surfaces and the interaction with other molecules,so the adsorption of molecules onto surfaces,and the interaction between molecules can be evaluated based on the change of surface charge.This dissertation focuses on fast and label-free monitoring of the interactions between macromolecules and surfaces or other materials with pulsed streaming potential(SP)measurement.With pulsed SP measurement,we quantitatively monitored the stability of polyelectrolyte layer,and the adsorption of protein onto bare capillary surface.The kinetic adsorption of nanoparticles onto polyelectrolyte layer and the interaction between graphene oxide(GO)and lysozyme(LYZ)have been monitored in situ with pulsed SP measurement.The dissertation consists of five chapters.Chapter I: Techniques that can be used to monitor the adsorption of molecules have been listed,especially SP method.The applications of SP measurement used in biological and chemistry,especially the interaction of proteins,polyelectrolytes,and NPs studied by SP measurement have been reviewed.Chapter II: The stability of polyelectrolytes varies significantly due to the properties of both polyelectrolytes and surfaces.Fast and reliable evaluation of the stability of the assembled layers is essential to optimize the assembling processes and select the best working conditions.In this work,the applicability of the pulsed SP measurement for real-time evaluation of the stability of assembled layers was monitored through surface charge.Polyethyleneimine(PEI)with a molecular weight of 1800 was selected as the model polyelectrolyte and the results confirmed the dependence of the stability of the assembled layers on the assembling conditions.A parameter SR,which is defined as the relative zeta potential change rate,was adopted to reflect the difference of the SP value of positively charged layer.Effect of assembling parameters,including pH,polyelectrolyte concentration,and contact time on the change of the SP were evaluated based on SR values.The feasibility of evaluation the stability of adsorbed proteins,oligonucleotide and cells on the formed PEI layer was also investigated by pulsed SP measurement and its reliability was confirmed by capillary electrophoresis.Chapter III: The adsorption of proteins onto the inner surface of capillary creates a major obstacle in all applications of capillary electrophoresis including the separation of proteins.It is necessary to evaluate the adsorption of proteins to the inner capillary surface with label-free and fast method.In this experiment,the adsorption of proteins was monitored with pulsed SP method,influence parameters such as the composition of buffer,buffer pH,metal ions,and the concentration of metal ions have been studied based on the initial SP change rate.Chapter IV: The deposition kinetics of GO onto PEI layer was characterized in situ with pulsed SP measurement,and it was found that the initial rate constant(ki)was dependent on the size of GO with same surface charge density at a fixed concentration under controlled experimental conditions.Assuming the deposition was controlled by diffusion at the initial stage,ki is proportional to Rh-2/3,where Rh is the hydrodynamic radius.By flushing a GO solution through a capillary coated with PEI layer,the initial change rate of relative SP(dEr/dt)was obtained in 20 s and ki was measured with five different concentrations in about 2 min.Three GO samples of different sizes obtained from the same batch of raw material were characterized with pulsed SP measurement to get ki values,and their sizes were verified with atomic force microscopy(AFM)and dynamic light scattering(DLS).The experimental results are consistent with the predicted effects of the size of NPs on their deposition kinetics.Chapter V: The adsorption of monodisperse silica NPs on the surface of capillary modified with PEI layer has been studied by pulsed SP measurement.The initial rate constant of silica NPs adsorbed onto PEI layer was calculated with five concentrations.By fitting the relative SP changes along with time of silica NPs onto PEI layer,the adsorption of silica NPs onto PEI layer was proved under diffusion controlled transport.For different sized silica NPs,the initial adsorption rate decreases with increasing particle size,which is correlated with the size of NPs(which got from DLS and scanning electron microscopy(TEM)method)and initial rate constant,pulsed SP may be used to monitor the size of spherical NPs.Chapter VI: Kinetic elucidation NP-protein interaction can facilitate the promoting applications of NPs in the biomedical fields and can be used as a kind of indicator of the nanosafety of NPs in biophysical conditions.In this experiment,the interaction between GO-LYZ has been monitored with pulsed SP measurement based on the charge quenching of GO and LYZ.With Pulsed SP measurement,the free concentration of LYZ in GO-LYZ solution can be attained,and the relationship between SP with constant KD and Hill coefficients n can be calculated.Impacts on GO-LYZ interaction from the incubation buffers(pH,conductivity,and salt solution)were evaluated on the basis of parameters KD and n values to interpret the interaction driving forces.Our study demonstrated the high simplicity and flexibility of pulsed SP measurement in probing the interaction between protein and NPs.Its reliability was confirmed by fluorescence spectrometry method.This method should be a rapid,label-free,and high throughput way method to measure the interaction between NPs with proteins,and could also be used to evaluate the effect of NPs on the adsorption kinetics of proteins and other bioamterials.