Point-of-Care Quantitative Measure of Glucose-6-Phosphate Dehydrogenase Enzyme Deficiency.

BACKGROUND AND OBJECTIVES: Widespread newborn screening on a point-of-care basis could prevent bilirubin neurotoxicity in newborns with glucose-6-phosphate dehydrogenase (G6PD) deficiency. We evaluated a quantitative G6PD assay on a digital microfluidic platform by comparing its performance with standard clinical methods. METHODS: G6PD activity was measured quantitatively by using digital microfluidic fluorescence and the gold standard fluorescence biochemical test on a convenience sample of 98 discarded blood samples. Twenty-four samples were designated as G6PD deficient. RESULTS: Mean ± SD G6PD activity for normal samples using the digital microfluidic method and the standard method, respectively, was 9.7 ± 2.8 and 11.1 ± 3.0 U/g hemoglobin (Hb), respectively; for G6PD-deficient samples, it was 0.8 ± 0.7 and 1.4 ± 0.9 U/g Hb. Bland-Altman analysis determined a mean difference of -0.96 ± 1.8 U/g Hb between the digital microfluidic fluorescence results and the standard biochemical test results. The lower and upper limits for the digital microfluidic platform were 4.5 to 19.5 U/g Hb for normal samples and 0.2 to 3.7 U/g Hb for G6PD-deficient samples. The lower and upper limits for the Stanford method were 5.5 to 20.7 U/g Hb for normal samples and 0.1 to 2.8 U/g Hb for G6PD-deficient samples. The measured activity discriminated between G6PD-deficient samples and normal samples with no overlap. CONCLUSIONS: Pending further validation, a digital microfluidics platform could be an accurate point-of-care screening tool for rapid newborn G6PD screening.

The dynamics and stability of lubricating oil films during droplet transport by electrowetting in microfluidic devices.

The operation of digital microfluidic devices with water droplets manipulated by electrowetting is critically dependent on the static and dynamic stability and lubrication properties of the oil films that separate the droplets from the solid surfaces. The factors determining the stability of the films and preventing surface fouling in such systems are not yet thoroughly understood and were experimentally investigated in this study. The experiments were performed using a standard digital microfluidic cartridge in which water droplets enclosed in a thin, oil-filled gap were transported over an array of electrodes. Stable, continuous oil films separated the droplets from the surfaces when the droplets were stationary. During droplet transport, capillary waves formed in the films on the electrode surfaces as the oil menisci receded. The waves evolved into dome-shaped oil lenses. Droplet deformation and oil displacement caused the films at the surface opposite the electrode array to transform into dimples of oil trapped over the centers of the droplets. Lower actuation voltages were associated with slower film thinning and formation of fewer, but larger, oil lenses. Lower ac frequencies induced oscillations in the droplets that caused the films to rupture. Films were also destabilized by addition of surfactants to the oil or droplet phases. Such a comprehensive understanding of the oil film behavior will enable more robust electrowetting-actuated lab-on-a-chip devices through prevention of loss of species from droplets and contamination of surfaces at points where films may break.

Novel application of digital microfluidics for the detection of biotinidase deficiency in newborns.

OBJECTIVE: Newborn screening for biotinidase deficiency can be performed using a fluorometric enzyme assay on dried blood spot specimens. As a pre-requisite to the consolidation of different enzymatic assays onto a single platform, we describe here a novel analytical method for detecting biotinidase deficiency using the same digital microfluidic cartridge that has already been demonstrated to screen for five lysosomal storage diseases (Pompe, Fabry, Gaucher, Hurler and Hunter) in a multiplex format. METHODS: A novel assay to quantify biotinidase concentration in dried blood spots (DBS) was developed and optimized on the digital microfluidic platform using proficiency testing samples from the Centers for Disease Control and Prevention. The enzymatic assay uses 4-methylumbelliferyl biotin as the fluorogenic substrate. Biotinidase deficiency assays were performed on normal (n=200) and deficient (n=7) newborn DBS specimens. RESULTS: Enzymatic activity analysis of biotinidase deficiency revealed distinct separation between normal and affected DBS specimens using digital microfluidics and these results matched the expected activity. CONCLUSIONS: This study has demonstrated performance of biotinidase deficiency assays by measurement of 4-methylumbelliferyl product on a digital microfluidic platform. Due to the inherent ease in multiplexing on such a platform, consolidation of other fluorometric assays onto a single cartridge may be realized.

Electric-field-controlled flow in nanoscale-thin wetting films.

A novel nanofluidic system based on electroosmotic flow in nanoscale-thin aqueous wetting films is reported. The water films formed spontaneously on mica substrates in a saturation humidity environment. The film thickness was determined to be a few tens of nanometers by optical interference and fluorescence intensity measurements and was consistent with a theoretical evaluation of the thickness of a film based on the competing forces of electrostatic repulsion and capillary pressure. Lateral flow was induced by applying a dc electric field tangential to the film and characterized by tracking the position of a fluorescent probe. The mobilities of the thin fluid layer and the flow marker were lower than the predictions of the electrokinetic theory, which may be a result of adsorption of the fluorescent molecules to the mica. Confinement of the film to two-dimensional "channels" was achieved by microcontact printing of patterned hydrophobic monolayers onto the substrate. This system has the advantage of simple and inexpensive fabrication in comparison to nanofluidic devices made by traditional lithography techniques.

Electric-field-assisted convective assembly of colloidal crystal coatings.

A new technique that combines evaporative convective deposition of colloidal crystal coatings with an electric field to achieve more rapid assembly and reduce the defects in the crystal structure is reported. When an ac voltage is applied across the particle suspension and the substrate in the convective assembly process, a longer film spreads from the meniscus by the electrowetting-on-dielectric (EWOD) effect. The data suggest that the EWOD-increased liquid surface area results in increased evaporation-driven particle flux and crystal assembly that is up to five times more rapid. The extended drying film also provides more time for particle rearrangement before the structure becomes fixed, resulting in formation of crystal domains an order of magnitude larger than those deposited by convective assembly alone. The results demonstrate that EWOD is a facile tool for controlling particle assembly processes in wetting films. The technique could be used in improved large-scale colloidal crystal coating processes.