This study proposed a method for the fine alignment of polydimethylsiloxane (PDMS) stamp used in multiple microcontact printing. The procedure of alignment method was organized into three steps; (1) the fabrication of PDMS stamp with an alignment marker, (2) the position detection of PDMS stamp with a noncontact laser displacement sensor on an automated planar x-y axes stage, and (3) the position adjustment of PDMS stamp to target position with another automated planar x-y-θ axes stage. The procedure was able to perform without contact onto the surface of PDMS stamp just before the moment of contact printing. Therefore, the surface would be safe from molecular/bacterial contamination. This study implemented the procedure into a developed system for the alignment of PDMS stamp. After adjusting the position of PDMS stamps applied with fibronectin-binding-fluorescent dyes with three different colors, contact printing was performed. As a result, the error of position between two different patterns of fibronectin was within 33 μm. This method would be useful for generating a multi-type-cellular tissue in tissue engineering and regenerative medicine.

Micro contact printing is a quite useful technique for patterning protein such as cellular adhesion molecular. Especially, the method is popular to fabricate substrates for cell patterning in tissue engineering. However, a manual printing is difficult, because the stamp made from polydimethylsiloxane (PDMS) is easily deformed, and a printed pattern is also crushed. This study focused on the stiffness of PDMS stamp and discussed a micro-contact-printing controlled stamping force for a higher quality of pattern than that of manual. Considering in availability in medical or biological laboratory, the measurement method of stamp stiffness was developed by using a general microscope. The proposed stamping method gave a high printing-quality with 2.5% error of stamping area. Stamping setup controlling stamping force was developed, and a protein was patterned on a cell-culture substrate by the setup. By using the protein-patterned substrate, two-cell-patterned co-culture was successfully performed.