Abstract
The ability to isolate specific, viable cell populations from mixed ensembles with minimal manipulation and within intra-operative time would provide significant advantages for autologous, cell-based therapies in regenerative medicine. Current cell-enrichment technologies are either slow, lack specificity and/or require labelling. Thus a rapid, label-free separation technology that does not affect cell functionality, viability or phenotype is highly desirable. Here, we demonstrate separation of viable from non-viable human stromal cells using remote dielectrophoresis, in which an electric field is coupled into a microfluidic channel using shear-horizontal surface acoustic waves, producing an array of virtual electrodes within the channel. This allows high-throughput dielectrophoretic cell separation in high conductivity, physiological-like fluids, overcoming the limitations of conventional dielectrophoresis. We demonstrate viable/non-viable separation efficacy of >98% in pre-purified mesenchymal stromal cells, extracted from human dental pulp, with no adverse effects on cell viability, or on their subsequent osteogenic capabilities.
Highlights
Separation and enrichment of cells derived from bone marrow, fat or even surgical waste are essential for a growing number of modern surgical cell-based therapies including in the treatment of diabetes[1], in vascular surgery[2] and in treatments for musculoskeletal disease[3]
The pressure waves are generated using a type of surface acoustic wave (SAW) commonly referred to as a Rayleigh-SAW (RSAW), in which a surface-propagating mechanical wave is established in a piezoelectric material by application of high-frequency signals to interdigitated transducers formed on the substrate surface
The device insertion loss was assessed as a function of buffer conductivity using a 2-port network analyser, and was found to be constant at −2 0.5 ± 0.5 dB across six orders of magnitude in conductivity (0.0001–10 Sm−1), indicating that even in high conductivities, no acoustic energy is dissipated into the fluidic system through, for example, Joule heating (Supplementary Fig. S1)
Summary
Separation and enrichment of cells derived from bone marrow, fat or even surgical waste (e.g. aspirate) are essential for a growing number of modern surgical cell-based therapies including in the treatment of diabetes[1], in vascular surgery[2] and in treatments for musculoskeletal disease[3]. Virtual electrodes are formed, with a pitch and periodicity equivalent to those of the externally located IDTs, but which cannot constitute an electron source, and electrochemical reactions (e.g. oxidation, reduction and radical formation), which lead to electrode fouling, bubble formation, heating, cell damage, pH gradients, inter alia, are prevented This means that cell separation is not limited to low-conductivity medium and can be achieved in conductivities >1 Sm−1 if required. The cells are only acted upon by DEP forces, causing them to align at regions of high and low field gradient, depending primarily on the relative polarizability of the cells and medium; SAW-DEP selectivity is optimised through careful choice of medium conductivity and SAW operating frequency, allowing their relative polarizabilities to be tuned This technique combines the high-throughput capabilities of acoustophoresis, with the exquisite discriminatory capabilities of DEP, and offers a possible solution to a wide range of current cell-separation challenges
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