0��10-9 g?mL-l. A 1��10-2 mol?L-l stock solution of potassium ferricyanide was prepared by dissolving the required amount of compound in distilled water. Diluted solutions of potassium ferricyanide were prepared by mixing portions of the stock solution with the required amounts of NaOH/NaHCO3 solution.2.2. Microchip FabricationPDMS is useful for microfluidic fabrication and application because of several important properties, including its mechanical flexibility, gas permeability and optical transparency. Moreover, its elasticity and hermetic self-sealing properties make multilayer construction of PDMS-based devices relatively straightforward. PDMS molding is used almost exclusively for rapid prototyping in corporate environments because of its simplicity and fast turnaround time. In this paper, the microchips employed in the experiments were fabricated with PDMS and glass using standard techniques. The PDMS chip is cured, removed, and then pressed or bonded onto a glass substrate to create a complex microchannel by the monolithic pouring method. The bottom chip was fabricated in glass using conventional wet chemical etching. In a final step, a glass chip was bonded to the PDMS chip to yield a network of closed channels. The upper layer and the bottom layer having different channels. The upper layer had the preconcentration microchannels (AB, 5 mm length and 150 ��m diameter) and the lower PDMS layer had ��Y�� shape reaction microchannels (150 ��m wide and 150 ��m deep). The reservoirs (R1, R2 R3 and R4) were punched on the upper PDMS layer using a round hollow punch,2.3. Monolith column polymerizationFor the polymerization of the monolith column butyl methacrylate (BMA), ethylene dimethacrylate (EDMA), 2,2��-azobis-(2-methylpropionitrile) and methanol/ethanol were used as the monomer, cross-linking agent, initiator and porogenic solvents, respectively. The detailed monolith column polymerization processes was carried out as follows: first, the microchannels flushed by sodium hydroxide solution and double distilled water were washed with silanization solution containing 30% (V/V) ��-MAPS in acetone. Prior to mixing with the porogenic solvents, EDMA and BMA were mixed with fresh basic alumina powder to remove the added contain inhibitor. After purging with nitrogen for 3 min, the mixture consisting of 75% porogenic solvents and 25% monomers were pumped into microchannel for the subsequent polymerization. The porogenic solvent used was a mixture of methanol and ethanol with a ratio of 5:3. A 1:1 mixture of BMA and EDMA was used as monomer. Then, 365 nm UV light was used to irradiate the mixture from the top from a 5 cm distance through a mask to control the location of the BMA monolithic column on the microchip. The BMA monolithic column was polymerized in-situ in an ultraviolet transparent PDMS microchannel on a homemade microfluidic chip; it was finally washed with methanol at 2.0 ��L?min-1 for 30 min.