Prepararion Of Optical Immunological Probe By Mesoporous Silica

1.A chemiluminescent signal amplification label (name as HRP-Ab2/SiO2) was proposed by co-immobilizing horseradish peroxidase (HRP) and HRP-labeled secondary antibody (Ab2) onto the mesoporous SiO2 nanoparticles. Based on a sandwich immunoassay, the immobilized monoclonal antibody ofα-fetoprotein (AFP) was used as a primary antibody, while HRP-Ab2/SiO2 was used as a secondary antibody. Due to the larger amount of HRP and Ab2 coated on the mesoporous SiO2, the HRP-Ab2/SiO2 labels could amplify the chemiluminescent signal of the system of luminol and H2O2. Under the optimal conditions, two linear ranges for AFP were obtained from 0.01 to 0.5 ng mL-1 and 0.5 to 100 ng mL-1 with a detection limit of 0.005 ng mL-1 (3σ). The proposed signal amplification strategy showed an excellent promise for sensitive detection of AFP and other tumor markers.2. A novel label-free chemiluminescent immunological probe was proposed based on the controlled release of medicine strategy. The immunological probe was prepared by immobilizing luminol molecular inside the mesoporous channels and immobilizing AFP antibody on the channel ends. Because all of the Si-OH groups on the external surface of MPS were terminated by chloro-silane, the random adsorption of protein or substrate reduced largely. Thus, the controlled release of luminol was confined inside the mesopores by changing the pH value of the solution. Based on one-step immunological reaction, the formed immunoconjugates blocked the pores, therefore the release of luminol from the mesopores was hindered and the changed CL signal was recorded for the detection of AFP. The fabricated label-free probe shows appropriate accuracy and offers an alternative to the multianalyte detection of antigens or other bioactive molecules.3. A novel parallel double-channel immunoassay strategy for the detection of two tumor markers: AFP and carcinoembryonic antigen (CEA) was proposed based on mesoporous magnetic materials (Fe3O4/SBA-15). Monoclonal antibodies of CEA and AFP were coated with HRP respectively on Fe3O4/SBA-15 mesoporous materials and used as the prime antibody. Antigens of CEA and AFP were coated with luminol on the gold nanoparticles (Au NPs) and used as the labeled antigen. Based on a competitive immunoassay format, sample antigen and labeled antigen competitively reacted to the limited active sites of the immobilized antibody. Then, immuno-complexes were injected into the flow cells, and were absorbed on the inside wall by the bar magnet. Through the control of two switches on two channels, H2O2 solution was injected into two different flow cells orderly, and different CL signals from two flow cells were sequentially detected. Under the optimal conditions, CEA and AFP could be assayed in the ranges of 1.0~80 ng mL?1 and 1.0~100 ng mL?1 with a detection limits of 0.2 ng mL?1.4. A novel ultra-sensitive immunoassay probe was proposed based on fluorescein isothiocyanate (FITC) labeled antibody modified mesoporous materials. The prepared signal amplification label has been successfully used to detect the level ofα-fetoprotein (AFP) in human serum samples. The sensitive fluorescent antibody modified mesoporous SBA-15 (FITC-Ab*/SBA-15) was prepared by covalent-linking FITC-labeled antibody onto the mesoporous SBA-15 surface by the EDC/NHS. AFP monoclonal antibodies were embedded into the glass sheet modified with Au-NP /mesoporous SBA-15. Based on a sandwich immunoassay format combined with flow injection analysis, two mixtures of the sample antigens and corresponding FITC-Ab*/SBA-15 were introduced into the flow cell for on-line incubation. After the sandwich immunoassay, FITC-Ab*/SBA-15was retained in the flow cell, and the fluorescent signal was recorded. More FITC-labeled antibody can be loaded base on the high surface area and large pore volume of mesoporous SBA-15 nanoparticles. Thus, the prepared amplification label can greatly improve the fluorescent response. The prepared amplification label possesses good bioactivity, comparable detection limit and linear range, and acceptable storage stability.