(786f) Nanoporous Membrane Molecular Sensors Based On Ion Currents

We report a new molecular (nucleic acid and protein) microfluidic sensor platform that is economical to mass produce, label-free,  multi-target (>5), rapid (<10 min), selective (SNP) and sensitive (pM)----and is hence most suitable for portable point-of-care diagnostic devices.  This new sensing platform employs fabricated or synthesized nanoporous membranes in PDMS, glass or polymer chips.  The synthesis techniques include solgel crystallization and photocuring.   When a DC field is applied across a nanoporous ion-selective membrane (or a fabricated nano-channel or nanostructure assembly),  a molecular-sized charge polarized layer develops on the depletion side after concentration polarization.  The presence of target molecules in this polarized layer can hence sensitively change the limiting and overlimiting currents of the membrane.  The overlimiting current shift is particularly large (~1 V) because large nucleic acids can change the hydrodynamic slip length and hence suppress the surface electrokinetic vortices responsible for the overlimiting current.   Oppositely charged small molecules, on the other hand, can produce a bipolar membrane like a PN semiconductor junction that can produce a large water-splitting ion current signal when a high field develops at the carrier-depleted junction with a reverse bias.  Without using electron transfer to detect the molecules, we eliminate the need for SAM and prevent any robustness issue due to spurious reactions, as in classical electrochemical sensing platforms.  This new molecular sensing technology also reduces the transport-limited assay time by several orders of magnitude because the high field in the polarized layer can attract large nucleic acids by dielectrophoresis and the depletion region can concentrate smaller ones in a flowing stream at the membrane sensor location.   We also use flow to linearize hairpin structures/collapsed globules and remove non-specifically bound molecules.   An optimum shear rate is found empirically as a function of the target molecule size and the probe-target association constant-----and is accurately captured by an activated theory for hybridization.  Since each membrane is only a micron-wide strip, extension to more than 10 targets can be easily done.   We have successfully tested and quantified this platform in double blind tests for RNAs of 4 Dengue virus seroltypes, microRNA of ovarian cancer cells, cytokine and interferon-gamma signal proteins,  Enterococcus, E coli, Brucella, Listeria, MRSA, Pseudomonas, Meningitis and four aquatic invasive species: quagga mussel, D bugenesis, E sinensis, L fortunei.  RNA detection is particularly exciting because of microRNA as a new cancer biomarker and the fact that many bacteria have tens of thousands of mRNA but only one copy of DNA implies that PCR amplification is not necessary even with only one pathogen per sample.  Prototype devices based on this platform is being developed and commercialized.  We also describe current extension of this sensor platform to massively parallel target detection and regenerable real-time sensors for immune cell cultures and remote applications.

Yossifon, G. and Chang, H.-C., “Selection of Nonequilibrium Overlimiting Currents: Universal Depletion Layer Formation Dynamics and Vortex Instability”, Phys Rev Lett., 101, 254501(2008).
Chang, H.-C. and Yossifon, G., "Understanding Electrokineitcs at the Nanoscale--a  Perspective", Biomicrofluidics, 3, 012001 (2009).
Basuray, S. , Senapati,  S.,  Ajian, A. , Mahon, A. R. and Chang, H.-C., “Shear and AC  Field Enhanced Carbon Nanotube Impedance Assay for Rapid, Sensitive and Mismatch-Discriminating DNA Hybridization”,  ACS Nano, 3, 1823 (2009).
Cheng, I.-F., S. Senapati, X. Cheng, S. Basuray, H-C Chang and H.-C. Chang, “A Rapid Field-Use Assay for Mismatch Number and Location of Hybridized DNAs”, Lab-on-a-Chip,10, 828-831 (2010).
Mahon, A. R., Barnes, M. A., Senapati, S., Feder, J., Chang, H.-C. and Lodge, D. M.,  “Molecular Detection of Invasive Species in Heterogeneous Mixtures using a Carbon Nanotube Platform”, PLOS One, 6, 17280 (2011).
Senapati, S., Basuray, S., Slouka, Z., Cheng, L.-J. and Chang, H.-C., "A Nanomembrane Based Nucleic Acid Sensing Platform for Portable Diagnostics" ,  Topics in Current Chemistry, 304, , 153-169(2011).
Chang, H.-C., Yossifon, G. and Demekhin, E. A. , “Nanoscale Electrokinetics and  Microvortices: How Microhydrodynamics Affects Nanofluidic Ion Flux”, Annual Rev of Fluid Mech, 44, 401-426 (2012).

Cheng, L.-J. and Chang, H.-C., “Microscale pH Actuation by Splitting Water”, Biomicrofluidics, 5, 046502 (2011).