We have demonstrated two types of size separation of biomolecules using a nanostructured matrix artificially fabricated using electron-beam lithography: sieve-type separation using a regular pillar array structure and size exclusion chromatography (SEC) type separation using a structure with narrow and wide gaps. With these devices, samples of double-stranded DNA molecules (2, 5, and 10 k base pairs) were clearly separated into bands; smaller molecules eluted earlier in the sieve type while they eluted later in the SEC type. The nanostructured matrix enables various types of molecular separation by changing the design of the nanostructure. Moreover, it should be easy to integrate the matrix with other biomolecular fluidic devices because it does not require a filling medium.

We have achieved the size separation of DNA molecules on a biochip using an anodic porous alumina as a separation matrix in electrophoresis. The bottom of the microfluid channel on the chip was made of a porous alumina membrane that has uniform nanoscale pores on its surface. Since smaller molecules are much more frequently trapped by nanopores than larger ones, larger molecules elute earlier than smaller molecules. This type of separation is the same process as that resulting from size-exclusion chromatography. When two kinds of DNA samples (3.2 and 0.3 kilobases) were applied to the biochip, they were clearly separated into two bands, with the larger molecules (3.2 kilobases) eluting earlier. Since nanopore formation does not require any nanolithography, the fabrication is easy and inexpensive. The matrix can be integrated with other biomicroelectromechanical systems reactors on a micro total analysis system chip.

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