Ultrathin Silicon Membranes for in Situ Optical Analysis of Nanoparticle Translocation across a Human Blood–Brain Barrier Model

Abstract

Here we present a blood-brain barrier (BBB) model that enables high-resolution imaging of nanoparticle (NP) interactions with endothelial cells and the capture of rare NP translocation events. The enabling technology is an ultrathin silicon nitride (SiN) membrane (0.5 μm pore size, 20% porosity, 400 nm thickness) integrated into a dual-chamber platform that facilitates imaging at low working distances (~50 μm). The platform, the μSiM-BBB (microfluidic Silicon Membrane-BBB), features human brain endothelial cells and primary astrocytes grown on opposite sides of the membrane. The human brain endothelial cells form tight junctions on the ultrathin membranes and exhibit a significantly higher resistance to FITC-dextran diffusion than *

Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells

“…The literature is lacking of 4 kDa Dextran permeability data on bEnd.3 cells and only one paper was found, the measured permeability coefficient was ranging from 2.48 to 4.01 x 10 −6 cm/s with a mean P e of 2.91 (0.43) x 10 −6 cm/s [ 34 ]. Our model was a bit leakier than this, as we measured a P e of 5.79 (0.44) x 10 −6 cm/s; this permeability value is similar to the hCMEC/D3 cell line, where the permeability of 4 kDa Dextran is ranging between ~5–14 x 10 −6 cm/s when cultured on Transwell membrane [ 35 – 37 ]. Our translocation studies suggest that GYR can transcytose across the BBB.…”

The Modular µSiM: a Mass Produced, Rapidly Assembled, and Reconfigurable Platform for the Study of Barrier Tissue ModelsIn Vitro

“…In this study, chips are oriented “trench-down” to culture a monolayer that is continuous across the window and non-window regions, however, higher resolution imaging can be achieved by flipping the chip into a “trench-up” orientation to bring the cells closer to the objective lens (Table S1, Supporting Information). [20]…”

“…The top of Component 2 is PSA that enables bonding to Component 1; 3) The third part is the membrane 'chip' which can be selected from nanoporous, microporous, or dual-scale (a mix of micro and nanopores) options based on application needs (Figure 1A, C). [18][19][20] Membrane chips have a trench side and flat side containing the membrane. [25] The membrane is free-standing over a window of 700 µm x 2 mm.…”

“…In this study, chips are oriented "trench-down" to culture a monolayer that is continuous across the window and non-window regions, however, higher resolution imaging can be achieved by flipping the chip into a "trench-up" orientation to bring the cells closer to the objective lens (Table S1, Supporting Information). [20] Assembly of the modular µSiM is a two-step process that utilizes PSA to irreversibly bond all components together (Video S1, Supporting Information). The PSA layers are protected by removable liners, which are added during component manufacture and removed just prior to m-µSiM assembly.…”

“…We have previously introduced the µSiM as a Transwell™-style culture microdevice featuring ultrathin (~100 nm), highly permeable, and optically clear silicon nitride membranes. [16][17][18] We have used the µSiM to create in vitro models of the blood-brain barrier, [19][20][21] the interface of the osteocyte lacuna-canalicular network in bone, [22][23][24] and as a tool to study leukocyte transmigration across vascular endothelium. [25][26] We addressed a need for high volume manufacturing and device reproducibility in a genetic screening of Staphylococcus aureus by enlisting a contract manufacturer, [22,24] but the approach produced large quantities of a configuration suitable to only one project.…”