Integrated serial dilution generation with a degassed centrifugal PDMS-based microfluidic device

A. Wang, K.W. Oh
University at Buffalo,
United States

Keywords: centrifugal microfluidic device, vacuum-assisted pumping

Summary:

Abstract This paper reports integrated serial dilution generations using a centrifugal force with a degassed PDMS made microfluidic device. Bio molecular and chemical concentration gradient is widely studied for the sample preparation and analysis. One of the successful serial dilution methods is using the centrifugal microfluidic with siphon valves [1]. However, the capillary force requires the surface modification to drive the sample. The vacuum-assisted force allows the device avoiding air bubbles as well as high tolerance in the surface condition [2]. Instead of the capillary force, we used the pre-degassed PDMS made device for using the vacuum-assisted force to convey samples from one chamber to the next dilution chamber. A microfluidic device for ten-fold dilution was proposed and evaluated with fluorescent dyes. Details The device schematic is shown in Fig. 1. The metering and dilution chambers with 640 um thickness each were fabricated using soft-lithography. The device was pre-degassed in a vacuum chamber before usage. The core operation principle is shown in Fig. 2. The phase-guide structure is the geometrical structure, which has slightly different height compared with other part of channels allowing confinement of samples within the structures. This phase-guide structures were used for assisting pre-determined volume transfer to the chamber. The sample transfer across the dilution chambers is based on vacuum-assisted force, which generates pressure difference across the liquid column. Sequential images of serial dilution and its procedure were shown in Fig. 3. First, the sample and buffer were loaded and filled into the metering chambers (Fig. 3 (a)), following the metering using vacuum-assisted force; 14.5 μl (#1) for the blue sample, 13.1 μl (#2, #3,#4) and 11.7 μl (#5) for the red buffer. (Fig. 3 (b)). Next, liquids were transferred to the dilution chambers by applying 1200 RPM (revolution per minute) (Fig. 3 (c)). Then, the excess amount of the samples (~1.4 μl, equivalent to 10% amount of sample in #1) into the adjacent dilution chamber using the vacuum-assisted force (Fig. 3 (d)). Next, the samples were mixed with the buffer by applying additional centrifugal force with changing acceleration rate over time. By repeating the procedure as shown in Fig. 3 (c) and Fig. 3 (d), the excess amount of the samples were kept transferred into the next dilution chamber (Fig. 3 (e), (f)) following the mixing with the buffer to complete the dilution. Serial dilution results were shown in the Fig. 4 (a). The fluorescent dye was used to evaluate reproducibility and quantitative dilution ratios (Fig. 4 (b)). Unfortunately, due to the background noise, the intensity in #5 chamber showed the negative value. Also, the intensity in #4 chamber was within the order of background noise, which could not be used for evaluation indicator. In conclusion, we proposed serial dilution methods utilizing centrifugal and vacuum-assisted force. The further evaluation is plan to be conducted such as volume analysis, Polymerase chain reaction (PCR) for solid proof-of-concept to evaluate further dilution ratios.