Nucleic acid extraction using a rapid, chemical free, ultrasonic technique for point-of-care diagnostics

Abstract

We have developed a miniature ultrasonic lysing system for the rapid release and extraction of Mycobacterium Tuberculosis (MTB) DNA. The prototype consists of two key sections, a disposable plastic manifold that hold sputum slides and a plastic cartridge with five fluidic DNA extraction channels. The MTB vaccine Bacillus Calmette-Guérin (BCG) was used as a simulant for MTB for all the lysing studies. The slide manifold houses up to five individual sputum slides and is processed by a computer to fill and load the cartridges with sputum suspensions from each slide. The plastic cartridge was developed to facilitate acoustic coupling through the bottom interface such that MTB/BCG can be lysed remotely. The disposable cartridge eliminated cross contamination problems and additional cleaning time. Due to the high power density, only 66 mW was required to lyse microliter BCG samples at the same level as the positive control (i.e. boiling for 30 minutes at 95 °C).

Rapid Nucleic Acid Extraction and Purification Using a Miniature Ultrasonic Technique

“…The details of the transducer operation and performance were cited previously [ 33 ]. Briefly, 36° Y-cut lithium niobate (90°, 90°, 36°) ( z, x, z ) excites a quasi-longitudinal wave with an electromechanical coupling of k t = 0.46 with an unloaded acoustic velocity of 7340 m/s.…”

“…The additional path length provided an additional tuning parameter between lysing efficacy and the subsequent nucleic acid extraction. For both interface types, Figure 4 b showed that increasing the applied power improved lysing efficiency, which required a balance between input power and heat generation by the transducer according to a thermal analysis [ 33 ].…”

“…The transducers were fabricated using 36° Y-cut lithium niobate (Boston Piezo-Optics, Bellingham, MA, USA), approximately 50 µm thick and 3 mm in diameter [ 33 ]. Gold (Au) electrodes were 0.2 µm thick with a 0.2 mm hold back from the edge of the substrates.…”

“…In our previous work, a low-power lysis approach was developed with negligible temperature rise in the microchannels for rapid extraction of Mycobacterium tuberculosis (MTB) DNA [ 33 ]. In this study, we investigated three types of DNA extraction methods integrated with a miniature bulk acoustic wave (BAW) transducer array on a disposable laminate device.…”

“…The electromechanical coupling ( k t 2 ) for 36° Y-cut lithium niobate is 46%, where the transducer operated from 50–70 MHz (see Appendix A ). A transmission line model and the finite-element method (FEM) were used to design the transducer and maximize acoustic coupling into disposable microchannels [ 33 ]. DNA was extracted and purified using three approaches: (1) silica-based sol-gel filled microchannels; (2) nucleic binding paramagnetic beads; and (3) Nafion-coated gold electrodes, where the extraction efficiency was compared for each method.…”