The Ligase Detection Reaction (LDR) is a mutation detection technique used to identify point mutations in deoxyribonucleic acid (DNA). A microscale Ligase Detection Reaction (LDR) device was designed and manufactured in polycarbonate. There are at least two mixing stages involved in the LDR identification process. Various micromixers were simulated in Fluent (v5.4, Lebanon, NH) and several test geometries were selected for fabrication. Passive diffusional micromixers were made with aspect ratios from 7 to 20. The mixers were made by SU-8 lithography, LIGA, laser ablation and micromilling to characterize each fabrication method. It was found that LIGA was best for making the micromixers, but was the longest process. The micromixers were fabricated and are being tested using fluorescent dyes. For a successful reaction temperatures of 0°C, 95°C and 65°C were needed. A stationary chamber method was used with thermal cycling in which the sample held while the temperature is cycled. Finite element analysis showed uniform temperatures in the rectangular 1.5 μl chambers and that air slits can effectively separate the thermal cycle zone from the 0°C cooling zone and the mixing region. A test device was laid out and micromilled with the temperature zones. A commercial thin film heater and a thermoelectric module were used with a PID controller to obtain the required process temperatures. Heating from 65°C to 95°C took 10 seconds, while cooling from 95°C to 65°C also took 10 seconds. The residence times at the required temperatures can adapt to changes in the LDR as parameters and reactant concentrations are varied.
Design of a Microfabricated Device for the Ligase Detection Reaction (LDR)
