Max M. Gong

Direct DNA Analysis with Paper-Based Ion Concentration Polarization

DNA analysis is essential for diagnosis and monitoring of many diseases. Conventional DNA testing is generally limited to the laboratory. Increasing access to relevant technologies can improve patient care and outcomes in both developed and developing regions. Here, we demonstrate direct DNA analysis in paper-based devices, uniquely enabled by ion concentration polarization at the interface of patterned nanoporous membranes in paper (paper-based ICP). Hepatitis B virus DNA targets in human serum are simultaneously preconcentrated, separated, and detected in a single 10 min operation. A limit of detection of 150 copies/mL is achieved without prior viral load amplification, sufficient for early diagnosis of hepatitis B. We clinically assess the DNA integrity of sperm cells in raw human semen samples. The percent DNA fragmentation results from the paper-based ICP devices strongly correlate (R(2) = 0.98) with the sperm chromatin structure assay. In all cases, agreement was 100% with respect to the clinical decision. Paper-based ICP can provide inexpensive and accessible advanced molecular diagnostics.

Nanoporous Membranes Enable Concentration and Transport in Fully Wet Paper-Based Assays

Low-cost paper-based assays are emerging as the platform for diagnostics worldwide. Paper does not, however, readily enable advanced functionality required for complex diagnostics, such as analyte concentration and controlled analyte transport. That is, after the initial wetting, no further analyte manipulation is possible. Here, we demonstrate active concentration and transport of analytes in fully wet paper-based assays by leveraging nanoporous material (mean pore diameter ≈ 4 nm) and ion concentration polarization. Two classes of devices are developed, an external stamp-like device with the nanoporous material separate from the paper-based assay, and an in-paper device patterned with the nanoporous material. Experimental results demonstrate up to 40-fold concentration of a fluorescent tracer in fully wet paper, and directional transport of the tracer over centimeters with efficiencies up to 96%. In-paper devices are applied to concentrate protein and colored dye, extending their limits of detection from ∼10 to ∼2 pmol/mL and from ∼40 to ∼10 μM, respectively. This approach is demonstrated in nitrocellulose membrane as well as paper, and the added cost of the nanoporous material is very low at ∼0.015 USD per device. The result is a major advance in analyte concentration and manipulation for the growing field of low-cost paper-based assays.

Turning the Page: Advancing Paper-Based Microfluidics for Broad Diagnostic Application

Infectious diseases are a major global health issue. Diagnosis is a critical first step in effectively managing their spread. Paper-based microfluidic diagnostics first emerged in 2007 as a low-cost alternative to conventional laboratory testing, with the goal of improving accessibility to medical diagnostics in developing countries. In this review, we examine the advances in paper-based microfluidic diagnostics for medical diagnosis in the context of global health from 2007 to 2016. The theory of fluid transport in paper is first presented. The next section examines the strategies that have been employed to control fluid and analyte transport in paper-based assays. Tasks such as mixing, timing, and sequential fluid delivery have been achieved in paper and have enabled analytical capabilities comparable to those of conventional laboratory methods. The following section examines paper-based sample processing and analysis. The most impactful advancement here has been the translation of nucleic acid analysis to a paper-based format. Smartphone-based analysis is another exciting development with potential for wide dissemination. The last core section of the review highlights emerging health applications, such as male fertility testing and wearable diagnostics. We conclude the review with the future outlook, remaining challenges, and emerging opportunities.

Hand-powered microfluidics: A membrane pump with a patient-to-chip syringe interface

In this paper, we present an on-chip hand-powered membrane pump using a robust patient-to-chip syringe interface. This approach enables safe sample collection, sample containment, integrated sharps disposal, high sample volume capacity, and controlled downstream flow with no electrical power requirements. Sample is manually injected into the device via a syringe and needle. The membrane pump inflates upon injection and subsequently deflates, delivering fluid to downstream components in a controlled manner. The device is fabricated from poly(methyl methacrylate) (PMMA) and silicone, using CO2 laser micromachining, with a total material cost of ∼0.20 USD/device. We experimentally demonstrate pump performance for both deionized (DI) water and undiluted, anticoagulated mouse whole blood, and characterize the behavior with reference to a resistor-capacitor electrical circuit analogy. Downstream output of the membrane pump is regulated, and scaled, by connecting multiple pumps in parallel. In contrast to existing on-chip pumping mechanisms that typically have low volume capacity (∼5 μL) and sample volume throughput (∼1-10 μl/min), the membrane pump offers high volume capacity (up to 240 μl) and sample volume throughput (up to 125 μl/min).

Paper-Based Quantification of Male Fertility Potential

BACKGROUND More than 70 million couples worldwide are affected by infertility, with male-factor infertility accounting for about half of the cases. Semen analysis is critical for determining male fertility potential, but conventional testing is costly and complex. Here, we demonstrate a paper-based microfluidic approach to quantify male fertility potential, simultaneously measuring 3 critical semen parameters in 10 min: live and motile sperm concentrations and sperm motility. METHODS The device measures the colorimetric change of yellow tetrazolium dye to purple formazan by the diaphorase flavoprotein enzyme present in metabolically active human sperm to quantify live and motile sperm concentration. Sperm motility was determined as the ratio of motile to live sperm. We assessed the performance of the device by use of clinical semen samples, in parallel with standard clinical approaches. RESULTS Detection limits of 8.46 and 15.18 million/mL were achieved for live and motile sperm concentrations, respectively. The live and motile sperm concentrations and motility values from our device correlated with those of the standard clinical approaches (R(2) ≥ 0.84). In all cases, our device provided 100% agreement in terms of clinical outcome. The device was also robust and could tolerate conditions of high absolute humidity (22.8 g/m(3)) up to 16 weeks when packaged with desiccant. CONCLUSIONS Our device outperforms existing commercial paper-based assays by quantitatively measuring live and motile sperm concentrations and motility, in only 10 min. This approach is applicable to current clinical practices as well as self-diagnostic applications.

Paper-based sperm DNA integrity analysis

We demonstrate a rapid, sensitive, and low cost paper-based approach for sperm chromatin integrity analysis that quantifies both DNA fragmentation and packaging. The paper-based test provides identical clinical outcome as the gold standard sperm chromatin structure assay. This technology is fast, simple, and suitable for broad application in clinics.