Fluorescent microparticles for sensing cell microenvironment oxygen levels within 3D scaffolds

Abstract

We present the development and characterization of fluorescent oxygen-sensing microparticles designed for measuring oxygen concentration in microenvironments existing within standard cell culture and transparent three-dimensional (3D) cell scaffolds. The microparticle synthesis employs poly(dimethylsiloxane) to encapsulate silica gel particles bound with an oxygen-sensitive luminophore as well as a reference or normalization fluorophore that is insensitive to oxygen. We developed a rapid, automated and non-invasive sensor analysis method based on fluorescence microscopy to measure oxygen concentration in a hydrogel scaffold. We demonstrate that the microparticles are non-cytotoxic and that their response is comparable to that of a traditional dissolved oxygen meter. Microparticle size (5–40 μm) was selected for microscale-mapping of oxygen concentration to allow measurements local to individual cells. Two methods of calibration were evaluated and revealed that the sensor system enables characterization of a range of hypoxic to hyperoxic conditions relevant to cell and tissue biology (i.e., pO2 10–160 mm Hg). The calibration analysis also revealed that the microparticles have a high fraction of quenched luminophore (0.90 ± 0.02), indicating that the reported approach provides significant advantages for sensor performance. This study thus reports a versatile oxygen-sensing technology that enables future correlations of local oxygen concentration with individual cell response in cultured engineered tissues.

Polydimethylsiloxane core–polycaprolactone shell nanofibers as biocompatible, real-time oxygen sensors

“…[40–42] The room temperature oxygen diffusivity in PDMS is 4.1×10 5 cm 2 /s, almost twice the diffusivity of oxygen in water (2.1×10 5 cm 2 /s) under the same conditions. [43] When used as sensor matrix in water, PDMS therefore provides relatively little barrier to oxygen diffusion to the probe and allows real-time reporting of oxygen levels. The recovery time is longer than the response time likely due to the fact that N 2 gas permeability and diffusivity in polymers is less than that of O 2 .…”

A novel microfluidic platform for high-resolution imaging of a three-dimensional cell culture under a controlled hypoxic environment

“…Zero pressure and convection flux conditions were set at media and gas outlets, and a no-slip condition was applied on the channel wall for the fluid flow analysis. The parameter values used for density, viscosity, and diffusivity of oxygen in media 27, 28, 30 , gel 34–36 , gas, and PDMS 26, 37, 38 are summarized in Table 1. In addition, diffusivity in the embedded film, D film , was assumed in a range of 2.0×10 −9 m 2 /s to 2.0×10 −12 m 2 /s, which corresponds to conditions in which the film is between two and 2,000 times less gas-permeable than PDMS, respectively.…”

A microfluidic device to study cancer metastasis under chronic and intermittent hypoxia

“…The oxygen-sensing microparticles were prepared as described by Acosta et al 36 However, for the particles presented here, no reference luminophore was employed. Briefly, 2 g of silica gel (Davisil, grade 710) was stirred in 40 ml of 0.01 N NaOH for 30 min.…”

“…The Ru(Ph 2 phen 3 )Cl 2 luminophore used in this study is a polar molecule and does not dissolve and distribute evenly in the PDMS film. Therefore, it was immobilized on a silica carrier that acts to maintain its charged, oxygen-sensing state, 36 and allows uniform distribution of the suspended particles within the 200 lm thick PDMS film below the cell-channel layer. Although this significantly modified the initial design, the high quantum yield and uniform distribution of these particles proved appropriate for accurate oxygen measurements and outweighed the disadvantage of including a 200 lm thick layer.…”