Fabrication of column chip made of PMMA for μFIA.

We proposed a low cost fabrication procedure of a poly(methylmethacrylate) (PMMA) column chip. 3D microchannel structure consisting of four columns in a chip for a mother die was fabricated using dry film photoresist and photolithography technique. Electroforming was applied to the mother die in order to obtain a Ni mold, then, the pattern was transferred to PMMA by hot press. The column had a dam structure to keep enzyme-immobilized microbeads with volume of 640 nL. The column chip was applied for a micro flow injection analysis (μFIA) system. For a demonstration, we measured lactose using two columns in series. One column was set on upper stream and filled with chitosan microbeads immobilized with β-galactosidase, the other was on downstream and filled with the beads immobilized with glucose oxidase. The lactose detection was accomplished less than 90s after the sample injection. The biosensing system also showed a high performance for lactose detection in wide range of 1 μM to 1mM. These results show that the column chip and our microfluidic biosensing system have the potential to assist minuaturization with small sample volume and short determination time for a sequential analysis.