Optimization of covalent antibody immobilization on macroporous silicon solid supports

In this paper, optimization of the protocol for covalent antibody immobilization on macroporous silicon solid supports of various porosities, which is recently being employed as a promising substrate for biosensors, has been reported. Covalent binding of antibody has been carried out by silanization and crosslinker attachment on the substrate. For maximum antibody immobilization on macroporous silicon, all the individual processes have been separately optimized for the first time in terms of treatment time, pH, concentration, incubation time and others with the help of optical density measurements. The optimum treatment of the surface after every step has been further reconfirmed by detailed EDX analysis, SEM measurements and contact angle measurements. It has been observed that the density of antibody binding increases with increasing porosity and for a 70% porosity macroporous sample it is almost three times more than that of planar silicon which is significantly higher than the previous comparative reports on planar silicon and macroporous silicon. The amount of properly oriented HIgG antibodies has been estimated by quantification of the alkaline phosphate conjugated protein A binding by optical density measurements.