Plasma membrane recovery kinetics of a microfluidic intracellular delivery platform

Abstract

Intracellular delivery of materials is a challenge in research and therapeutic applications. Physical methods of plasma membrane disruption have recently emerged as an approach to facilitate the delivery of a variety of macromolecules to a range of cell types. We use the microfluidic CellSqueeze delivery platform to examine the kinetics of plasma membrane recovery after disruption and its dependence on the calcium content of the surrounding buffer (~5min without calcium vs. ~30s with calcium). Moreover, we illustrate that manipulation of the membrane repair kinetics can yield up to 5x improvement in delivery efficiency without significantly impacting cell viability. Membrane repair characteristics initially observed in HeLa cells are shown to translate to primary naïve murine T cells. Subsequent manipulation of membrane repair kinetics also enables the delivery of larger materials, such as antibodies, to these difficult to manipulate cells. This work provides insight into the membrane repair process in response to mechanical delivery and could potentially enable the development of improved delivery methods.

A Size‐Selective Intracellular Delivery Platform

“…This cell suspension was mixed with a tetramethylrhodamine‐conjugated 10 kDa dextran fluorescent dye and passed through a chip whose constrictions were 7 μm wide ( Figure A). Tetramethylrhodamine‐conjugated dextran was chosen for these experiments for compatibility with downstream FACS analysis; while larger than the 3 kDa cascade blue‐conjugated dextran employed earlier, delivery efficiency is not affected by the molecular weight of dextran . GFP expression in PANC‐1 cells provided an independent signal to confirm successful labeling of tumor cells.…”

“…The microfluidic device has demonstrated applicability in delivering a variety of materials with a wide range of sizes that are retained by the cells, allowing for the manipulation of their biological properties . Here, our use of in vitro CTC models and clinical PDAC patient samples has demonstrated the additional utility of this platform to selectively deliver payload within a heterogeneous population of cells.…”

“…The delivery of a wide range of cargo sizes (3 kDa to 2 MDa) can be achieved with high efficiency and uniform distributions, suggesting that the size of holes resulting from the membrane disruption are homogeneous within that population of cells . While this transient disruption of the cell membrane allows for bidirectional movement of material across the membrane, cells remain viable and retain their proliferative capacity and biological activity . Here we demonstrate the use of a microfluidic system to confer selective cytosolic delivery within a mixture of cells based on cell size.…”

“…A method for cytosolic delivery that is independent of surface‐expressed markers on cells has been demonstrated using a microfluidic device that disrupts the cell membrane using mechanical forces as cells traverse through tight constrictions . The delivery of a wide range of cargo sizes (3 kDa to 2 MDa) can be achieved with high efficiency and uniform distributions, suggesting that the size of holes resulting from the membrane disruption are homogeneous within that population of cells . While this transient disruption of the cell membrane allows for bidirectional movement of material across the membrane, cells remain viable and retain their proliferative capacity and biological activity .…”

Cyclic Strain Enhances Cellular Uptake of Nanoparticles

“…Researchers reported that cyclic stretch affects the mechanical properties of ECs, such as cell proliferation [14], cytoskeletal structure [15], signal transduction [16], and gene expression and transfer [17]. However very few researches have been done to quantify the effect of cyclic stretching on the cellular uptake of NPs [18]. The aim of 2 Journal of Nanomaterials this study is to investigate the effects of cyclic stretch on the cellular uptake of NPs and its dependence on NP size through controlled tests.…”