Microfluidic System for Facilitated Quantification of Nanoparticle Accumulation to Cells Under Laminar Flow

Abstract

The identification of novel, synthetic targeting ligands to endothelial receptors has led to the rapid development of targeted nanoparticles for drug, gene and imaging probe delivery. Central to development and optimization are effective models for assessing particle binding in vitro. Here, we developed a simple and cost effective method to quantitatively assess nanoparticle accumulation under physiologically-relevant laminar flow. We designed reversibly vacuum–sealed PDMS microfluidic chambers compatible with 35 mm petri dishes, which deliver uniform or gradient shear stress. These chambers have sufficient surface area for facile cell collection for particle accumulation quantitation through FACS. We tested this model by synthesizing and flowing liposomes coated with APN (KD ~ 300 µM) and VCAM-1-targeting (KD ~ 30 µM) peptides over HUVEC. Particle binding significantly increased with ligand concentration (up to 6 mol%) and decreased with excess PEG. While the accumulation of particles with the lower affinity ligand decreased with shear, accumulation of those with the higher affinity ligand was highest in a low shear environment (2.4 dyne/cm2), as compared with greater shear or the absence of shear. We describe here a robust flow chamber model that is applied to optimize the properties of 100 nm liposomes targeted to inflamed endothelium.

Gold nanoparticle interactions with endothelial cells cultured under physiological conditions

“…27 Based on the cell viability determined in the various assays, NP concentrations of 12.5 to 25 µg / mL were used in further studies. The images presented in Figure 1 E 16,32 and enhanced the accumulation of liposomes targeted to vascular cell adhesion molecule-(VCAM-) 1 and also non-targeted nanoparticles in endothelial cells, 32,33 nanoparticle-cell interactions were explored under physiological medium flow conditions. The cartoon in Figure 2 A shows the movement of the added NPs across the cells under flow conditions.…”

Preliminary protein corona formation stabilizes gold nanoparticles and improves deposition efficiency

“…As standard in vitro models operate under static conditions, fluid dynamic variations to NP and cellular behavior are neglected in these models. Preliminary work has identified that the introduction of dynamic flow modified cellular morphology, NP behavior, and NP dosimetry (Breitner et al 2015;Kusunose et al 2013;Ucciferri et al 2014). These results suggest that the nano-cellular interface and the rate of NP deposition will be influenced by dynamic flow; making it a critical variable to include during AuNP application analysis.…”

Interactions of antitumour Sialyl Lewis X liposomes with vascular endothelial cells

“…Initially, relative binding avidity of HUVEC for targeted and nontargeted liposomes in the range of the ligand concentrations was explored according to [34]. Activated or non-activated cells were incubated with the formulations under endocytosis-blocking conditions (at 4°C); then, the cells were allowed to consume surface-bound liposomes (at 37°C) and were analysed by FACS.…”

“…Relative binding avidity was assessed according to [34] as follows: cells were chilled to 4°C, washed with cold DPBS and incubated for 1 h on ice with 2% SiaLe X or control liposomes (50 μM total lipid, 200 μL serum-free medium), then rinsed 3 times with DPBS, treated with 500 μL of complete medium and incubated for 1 h at 37°C. Cells were then rinsed with DPBS, detached with EDTA solution (10 min 37°C) and analysed by FACS.…”