Aliquoting on the centrifugal microfluidic platform based on centrifugo-pneumatic valves

We present a new method for aliquoting liquids on the centrifugal microfluidic platform. Aliquoting is an essential unit operation to perform multiple parallel assays (geometric multiplexing) from one individual sample, such as genotyping by real-time polymerase chain reactions (PCR), or homogeneous immunoassay panels. Our method is a two-stage process with an initial metering phase and a subsequent transport phase initiated by switching a centrifugo-pneumatic valve. The method enables aliquoting liquids into completely separated reaction cavities. It includes precise metering that is independent on the volume of pre-stored reagents in the receiving cavities. It further excludes any cross-contamination between the receiving cavities. We characterized the performance for prototypes fabricated by three different technologies: micro-milling, thermoforming of foils, and injection molding. An initial volume of similar to 90 mu l was split into 8 aliquots of 10 mu l volume each plus a waste reservoir on a thermoformed foil disk resulting in a coefficient of variation (CV) of the metered volumes of 3.6%. A similar volume of similar to 105 mu l was split into 16 aliquots of 6 mu l volume each on micro-milled and injection-molded disks and the corresponding CVs were 2.8 and 2.2%, respectively. Thus, the compatibility of the novel aliquoting structure to the aforementioned prototyping and production technologies is demonstrated. Additionally, the important question of achievable volume precision of the aliquoting structure with respect to the production tolerances inherent to each of these production technologies is addressed experimentally and theoretically. The new method is amenable to low cost mass production, since it does not require any post-replication surface modifications like hydrophobic patches.