Dual dielectropheretic membrane for monitoring cell migration
Inventors
Reyes-Hernandez, Darwin R. • NABLO, BRIAN
Assignees
United States Department of Commerce
Publication Number
US-10514359-B2
Publication Date
2019-12-24
Expiration Date
2032-09-21
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Abstract
A dual dielectropheretic article for monitoring cell migration includes: a membrane to selectively migrate a plurality of cells across the membrane, the membrane including: a first surface to receive the cells; a second surface opposed to the first surface; and a plurality of communication paths disposed in the membrane to provide the selective migration of the cells across the membrane from the first surface to the second surface; a first electrode disposed on the first surface to: provide an electric field for dielectrophoresis of the cells at the first surface; and provide a first potential for monitoring an impedance at the first surface; and a third electrode disposed on the second surface to: provide an electric field for dielectrophoresis of the cells at the second surface; and provide a third potential for monitoring an impedance at the second surface.
Core Innovation
The invention is a dual dielectropheretic article for monitoring cell migration comprising a membrane designed to selectively migrate cells from a first surface to a second surface. The membrane includes a plurality of communication paths and has electrodes disposed on both the first and second surfaces. These electrodes generate electric fields for dielectrophoresis of cells at each surface and provide potentials for monitoring impedance, thereby enabling real-time or quantitative measurements of cell migration and invasion through the porous membrane.
The invention overcomes the problem of efficiently capturing and monitoring cell migration in microfluidic devices while maintaining cell viability and function. The background discusses difficulties with existing dielectrophoretic systems in supporting long term cell adhesion, viability, and differentiation especially under low conductivity media and electric field exposure. It also addresses the challenge of monitoring selective migration of cells across membranes, the need for multilayer microfluidic devices enabling cell-cell interaction studies, and the limitations of mechanical trapping systems that disrupt flow conditions.
The disclosed article integrates microfabricated electrical components on opposing surfaces of a permeable membrane to produce electronic signals representing cell movement as cells traverse the membrane. The system also incorporates a hybrid cell adhesive material (hCAM) comprising layers of polyelectrolytes and fibronectin adapted for instantaneous cell adhesion post dielectrophoretic manipulation and supporting long term cell functions including proliferation and differentiation. The article allows dielectrophoretic trapping, electrical impedance monitoring on both membrane surfaces, and optional optical microscopy to monitor migration, all within a multilayer microfluidic device simulating physiological flow environments.
Claims Coverage
The patent contains 19 claims and includes multiple independent claims directed to the dual dielectropheretic article and a process for determining impedance. The claims cover inventive features related to membrane structure, electrode arrangement, fluidic integration, impedance monitoring, and cell adhesive materials.
Membrane with dual surface electrodes for dielectrophoresis and impedance monitoring
A membrane with first and second opposed surfaces, having a plurality of communication paths for selective cell migration; first and second electrodes disposed on the first surface to provide an electric field for dielectrophoresis and impedance monitoring; third and fourth electrodes on the second surface similarly providing dielectrophoresis and impedance monitoring.
Integration of membrane with substrate and dual flow channels
Membrane disposed on a substrate; a first flow channel on the substrate in fluid communication with the first membrane surface to provide cells for dielectrophoresis and migration; a second flow channel on the substrate in fluid communication with the second membrane surface to receive migrated cells.
Use of probe electrodes spatially separated from the membrane for impedance measurement
First and second probe electrodes disposed on the substrate, electrically communicating with the first and second flow channels respectively; these provide additional potentials for impedance monitoring in combination with respective electrodes on the membrane.
Impedance analyzer electrical configurations for monitoring migration
Electrical connection of impedance analyzer to membrane electrodes and probe electrodes enabling two-probe, three-probe, or four-probe impedance measurements at first and second membrane surfaces to monitor cell migration through impedance changes.
Incorporation of optical objective for imaging migration
An optical objective disposed near the second flow channel and membrane second surface to provide optical microscopy within the field of view for monitoring cell migration from the first to the second membrane surface.
Use of cell adhesive material on membrane surfaces
Cell adhesive material comprising extracellular matrix components and/or polyelectrolytes disposed on first surface, second surface, or both to adhere the cells and maintain viability during assays.
Membrane pore characteristics for selective migration
Communication paths comprising open pores with diameters ranging from 2 nm to 100 μm arranged in ordered or random patterns to allow selective migration of cells across the membrane.
Electrode material composition
First and third electrodes independently comprising metal, electrically conductive polymers, glass, ceramics, semiconductors, or combinations thereof.
Process for determining impedance in the dual dielectropheretic article
Providing the dual dielectropheretic article; applying alternating current voltages to electrodes or probe electrodes; monitoring electrical responses on electrodes not provisioned with the AC voltage; converting electrical responses to impedance values at the first and second membrane surfaces to reflect cell migration.
The independent claims encompass a multilayer microfluidic dual dielectropheretic article featuring a permeable membrane with electrode pairs on each side for both dielectrophoretic cell trapping and impedance-based migration monitoring, optionally using probe electrodes and optical microscopy, combined with cell adhesive material for maintaining cell viability and function, and a process utilizing this article to measure impedance changes correlating to cell movement.
Stated Advantages
Provides real-time or quantitative measurement of cell migration across a porous membrane using electrical impedance.
Allows dielectrophoretic trapping and selective control of cells on membrane surfaces for efficient cell enrichment.
Supports long term viability, adhesion, proliferation, and differentiation of cells under DEP conditions including low conductivity media and electric fields.
Integrates multiple functionalities (DEP trapping, impedance monitoring, and optical microscopy) within a multilayer microfluidic device simulating physiological conditions.
Enables normalized quantification of cell migration by having electrodes on both sides of the membrane within the same device.
Facilitates small sample and reagent volumes, better microenvironment control, and reduced waste through microfluidic integration.
Documented Applications
Monitoring of cell migration and invasion through a porous membrane in real-time for cell function studies.
Long term cell culture experiments followed by induced cell differentiation such as neuronal differentiation of P19 cells.
Cell-cell interaction studies in multilayer microfluidic co-culture systems with physical separation of different cell types.
Assessment of cell adhesive materials under dielectrophoretic trapping conditions and subsequent cell proliferation and differentiation.
In vitro studies of drug transport, cell monolayer permeability, and co-cultures combining dielectrophoresis and permeable membrane technology.
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