Fabricating arrays of single protein molecules on glass using microcontact printing

Microcontact printing biomolecules from elastomeric micropatterned stamps onto surfaces is a versatile method to prepare surfaces for diagnostic applications. We show how to create patterns of proteins having a lengthscale lower than 100 nm using high-resolution microcontact printing. The elastomeric stamps used have meshes composed of 100- and 40-nm-wide lines, arrays of 100 x 400 nm(2) features, and arrays of 100-nm-wide,posts. The spherical geometry of the posts on the stamps contributes to reduce the printed areas below the effective size of the molded features. Proteins adsorb onto the hydrophobic surface of the stamp during the inking step, and by varying the concentration of the protein solutions, it is possible to adsorb a single or a few protein molecules, such as antibodies (fluorescently labeled) or green fluorescence proteins, on each of the elements forming the high-resolution pattern of the stamp. The transfer of the proteins from the stamp to a hydrophilic glass surface occurs during the printing step. Characterization of the printed patterns using atomic force microscopy and fluorescence confocal microscopy reveals sites unoccupied or occupied by one or more protein molecules that are located within 50 nm of the expected printed locations. The placement of a small number of protein molecules on a surface at precise locations is the key to localizing and identifying single proteins and might constitute a method of choice to study single protein molecules on surfaces.