(J-366) Surface Acoustic Wave Microfluidics for Rapid Sickle Cell Disease Screening via Characterization of Blood Plasma Protein Concentration

Introduction:: Sickle cell disease (SCD) is a genetic blood disorder that affects millions of people worldwide. It is particularly prevalent in the developing world, where resource limitations create barriers for using conventional laboratory-based diagnostic methods. This prevents widespread screening and early treatment, leading to high childhood mortality. Recently, researchers have turned to microfluidic devices to provide ultraportable, rapid diagnostic methods that can be carried out in low resource settings. However, currently existing microfluidic approaches still suffer from complicated fabrication, requirements of skilled operators, or lack of sensitivity. Here, we introduce a microfluidic device that integrates surface acoustic waves (SAWs) to analyze relative total protein concentrations in blood plasma, which can be used as a rapid prescreening method for SCD.


Materials and Methods:: The SAW microfluidic device consists of a rectangular polydimethylsiloxane (PDMS) channel bonded to a piezoelectric lithium niobate substrate. The channel has a height of 80 µm and a width of 1 mm. A pair of identical interdigital transducers were fabricated on the substate on either side of the channel. Upon actuation via a radio frequency (RF) input (2 W, 20 MHz), SAWs propagate along the substrate towards the PDMS chamber. When the waves contact fluid in the channel, the interference from identical SAWs propagating towards each other create a patterned standing SAW field of high- and low-pressure nodes in the fluid. Additionally, the SAW energy induces rapid heating of the fluid up to 75 C within 90 seconds.

Whole blood samples were centrifuged at 2000 rcf for 10 minutes to separate the supernatant plasma from red blood cells, which was then used for analysis. After loading the plasma sample in the channel, the combination of acoustic patterning and heating caused proteins in the plasma to rapidly unfold and precipitate, while simultaneously concentrating and assembling along the nodal lines formed by the standing SAW. We tracked the precipitation process by plotting the change in grayscale intensity at the nodal lines to generate a protein melting curve. To quantify the relative protein concentration in the sample, we calculated the difference between the maximum and minimum grayscale intensity values on the curve.


Results, Conclusions, and Discussions:: We first compared the relative concentration difference between plasma samples from centrifuged whole blood taken from a healthy (HbA) and a post transfusion waste (HbSS - SCD) patient. We found that the HbSS sample exhibits a significantly higher relative difference in greyscale intensity at 90 seconds (approximately 75 C), suggesting a much higher relative concentration of protein compared to the HbA sample.

To confirm that the intensity difference stemmed from total protein concentration in the selected sample, we compared three different dilutions of plasma in phosphate-buffered saline (PBS). As we lowered the plasma content in the sample, we found that the difference in greyscale intensity at 75C also decreases significantly, demonstrating the sensitivity that our device provides. This also illustrates a generalized method that can be used to compare protein concentrations in any solution.

In our future work, we would like to gain a further understanding of which specific proteins in plasma provide the most significant signal in the protein melting curve. It is well known that a wide variety of proteins, particularly those associated with inflammation, are elevated in SCD. A targeted study to help pinpoint these proteins that show consistently elevated levels in the plasma of SCD patients will be completed. Furthermore, our device shows promise as a method to screen for the impact different types of SCD disorders, and possibly in contrast to SC trait, using a technique that is reliable and easy to apply in low-resource areas that are particularly difficult to reach using current standard diagnostic methods.


Acknowledgements (Optional): : The SAW microfluidic devices were fabricated in the JILA clean room at University of Colorado Boulder.

References (Optional): :