The Application Of Differential Streaming Potential Measurement For Protein Adsorption And Label-free Quantitative Analysis

Protein adsorption at liquid-solid interfaces has been a focus of much past research and continues to be a focus of research because of its importance to the development of biocompatible materials, diagnostic kits, and protein separations. Differential streaming potential measurement is a fast, sensitive and simple method to characterize the surface charge of microchannel. In the condition of certain pH and conductivity, the change of channel surface charge caused by protein adsorption can influence the streaming potential values of both ends of microchannel, therefore, the protein adsorption can be analysed indirectly by monitoring the change of streaming potential values. In addition, the streaming potential measurement can be used for quantitative determination of protein content in a solution, in this process, protein can maintain activity because it need not to be marked. It is believed that the streaming potential measurement has great application prospects in the field of protein analysis for less time-consuming and simpler procedures.This dissertation mainly focused on the application of differential streaming potential measurment for protein adsorption and label-free quantitative analysis, carrying out the studies as follow three aspects:1. A novel pulse differential streaming potential measurement device is built and its applicability is determined. On this basis, the adsorption behavior of four proteins on the fused silica capillary surface are studied, the binding capacity of proteins on surface is examined from two aspects, one is the time that streaming potential reached equilibrium value required, the other one is the difference values of streaming potential before and after protein layer flushed by buffer solution, the findings are appiled to the adsorption of proteins marked by FITC. The experiment test the adsorption behavior of lysozyme (LYZ) and bovine serum albumin (BSA) at different adsorption stages and make an explanation for "Overshoot" phenomenon in the LYZ adsorption process. At the same time, the adsorption rates are derived under the condition of different pHs, respectively. Finally, the relationship between streaming potential and adsorption amount is discussed. The results show that the streaming potential measurement is a simple and fast analysis technology with the advantages of less sample reagent consuming and wider application in the protein adsorption analysis field.2. On the basis of single protein adsorption studies, the studies of mixed proteins adsorption behavior on the capillary surface also are carried out. The results of proteins sequence adsorption and simultaneous adsorption show that replace phenomenon exist between BSA and bovine fibrinogen (BFG) and BSA can interact with LYZ on the capillary surface, the condition of protein A and immune globulin (IgG) is similar to BSA and LYZ. According to the derived mathematical model, the equilibrium constant of protein-protein interaction can be obtained. The results show that the simple and fast streaming potential measurement can be expected to become a new method to study protein interaction.3. Antigen NSE is immobilized on the aminosilane surface with glutaraldehyde successfully. The final modified surface can prevent the nonspecific adsorption of proteins by coating the polyvinyl alcohol (PVA) and double blocking with BSA. On this basis, the standard curve of anti-NSE with other interference proteins as many as2×102～2×105times content of anti-NSE is determined in the range of10-900ng/mL. At last, the test results of a healthy blood sample indicate that the normal content of anti-NSE is about160ng/mL.