Binding of labeled antigens to antibody arrays allows for the simultaneous analysis of a wide variety of proteins. One challenge in using such arrays is that the concentrations of some species such as hormones can be as low as 10-12 g/mL. In an effort to enhance the sensitivity of analysis with antibody arrays, we began developing arrays in porous alumina membranes. Because the membranes are optically transparent and have an internal surface area that is 500-fold greater than that of typical glass substrates, arrays in porous alumina have the potential to yield a 500-fold increase in sensitivity relative to traditional arrays. Although a few previous studies showed the potential of membranes in such analyses, our procedure for forming the arrays, which is shown below, is especially simple and avoids the need for blocking to reduce nonspecific adsorption. Layer-by-layer adsorption of a polyelectrolyte multilayer containing poly(acrylic acid) (PAA) and protonated poly(allylamine) (PAH) followed by activation of the carboxylic acids of PAA to N-hydroxysuccinimide esters yields a surface that covalently binds antibodies by reaction with their amino groups. Anti-IgG arrays prepared using these membranes show a 500-fold decrease in dection limit as expected. For more details on the analysis, click here.