Screening Estrogen Receptor Modulators in a Paper-Based Breast Cancer Model

Abstract

The health risks associated with acute and prolonged exposure to estrogen receptor (ER) modulators has led to a concerted effort to identify and prioritize potential disruptors present in the environment. ER agonists and antagonists are identified with end-point assays, quantifying changes in cellular proliferation or gene transactivation in monolayers of estrogen receptor alpha expressing (ER+) cells upon exposure. While these monolayer cultures can be prepared, dosed, and analyzed in a highly parallelized manner, they are unable to predict the potencies of ER modulators in vivo accurately. Physiologically relevant model systems that better predict tissue- or organ-level responses are needed. To address this need, we describe here a screening platform capable of quantitatively assessing ER modulators in 96 chemically isolated 3D cultures. These cultures are supported in wax-patterned paper scaffolds whose design has improved performance and throughput over previously described paper-based setups. To highlight the potential of paper-based cultures for toxicity screens, we measured the potency of known ER modulators with a luciferase-based reporter assay. We also quantified the proliferation and invasion of two ER+ cell lines in the presence of estradiol. Despite the inability of the current setup to better predict in vivo potencies of ER modulators than monolayer cultures, the results demonstrate the potential of this platform to support increasingly complex and physiologically relevant tissue-like structures for environmental chemical risk assessment.

Paper-based Transwell assays: an inexpensive alternative to study cellular invasion

“…We recently showed that 96-zone paper scaffolds could be loaded consistently with a multichannel pipette with RSD values of 10.6%–21.4% across a single deposition; 8.6%–19.9% across multiple depositions with the same pipette tip, and; 15.1% across an entire plate. 30 While reproducible seeding is user dependent and improves with practice, inconsistencies in cell density can be eliminated with the incorporation of liquid handlers capable of accurately and reproducibly seeding each zone.…”

Generating linear oxygen gradients across 3D cell cultures with block-layered oxygen controlled chips (BLOCCs)

“…Acrylic and food-grade silicone are biocompatible and have been used to prepare microfabricated devices for cell culture previously. [29][30][31] To ensure the laser-cut components † Electronic supplementary information (ESI) available. See DOI: 10.1039/c9ay01690b could support prolonged cell culture, we incubated them in contact with parental M231 cells suspended in Matrigel at a final density of 2000 cells per μL.…”

Fast Strategy to Functional Paper Surfaces

“…Owing to its significant advantages such as low-cost, easy fabrication and disposal, flexibility and renewability, paper has played a very important role in the propagation and development of human civilization since ancient times. − Various types of paper with different physical and chemical properties were created in the past thousands of years, making a huge library of substrate materials for both industry production and scientific research. The easy deformation properties (by cutting or folding) of the paper have also led to various innovative technologies and applications. − Because of these superiorities, paper has been widely applied in publications, packages, point-of-care test (POCT) devices, filters, and so forth, in every corner of our daily life. − It has also been frequently employed in scientific research, as substrate materials to grow nanomaterials, − perform bioanalysis, ,− culture cells, − and fabricate various paper-based devices. − …”

“…In many applications, paper has to be functionalized to alter its physical/chemical properties or bioactivities, for example, hydrophilicity, reactivity, biocompatibility, antifouling and antibacterial performances. , Patterning of physical and chemical properties on paper is also commonly required in the fabrication of paper-based microarrays and microfluidics, which are widely used in cell incubation devices, − POCT settings, , and paper-based electronic devices. , Many techniques have been used to functionalize paper and create patterns. Chemical functionalization on pure cellulose paper could easily be achieved because of the existence of numerous hydroxyl groups on cellulose molecules, which can react with various functional groups, allowing the covalent attachment of designed molecules on the paper substrate.…”

“…Many reported methods can only introduce alkyl or fluorinated chains onto paper substrates, which significantly limited the potential application of the functionalized paper substrates. For patterning on paper, methods such as wax printing, photolithography, micro-contact printing, and plasma treatment have been developed. ,,, However, these methods usually require time-consuming steps and specific devices, and most of them can only generate patterns with low resolution (>200 μm), far from the requirement for high-density cell arrays and precision microfluidic/electronic devices. A paper-functionalization method that is general (can be applied on different types of paper and allows various functional groups to be attached on paper substrates), highly efficient, low-cost, and patternable is highly demanded.…”