Abstract
Multicellular spheroids represent a well-established 3D model to study healthy and diseased cells in vitro. The use of conventional 3D cell culture platforms for the generation of multicellular spheroids is limited to cell types that easily self-assemble into spheroids because less adhesive cells fail to form stable aggregates. A high-precision micromoulding technique developed in our laboratory produces deep conical agarose microwell arrays that allow the cultivation of uniform multicellular aggregates, irrespective of the spheroid formation capacity of the cells. Such hydrogel arrays warrant a steady nutrient supply for several weeks, permit live volumetric measurements to monitor cell growth, enable immunohistochemical staining, fluorescence-based microscopy, and facilitate immediate harvesting of cell aggregates. This system also allows co-cultures of two distinct cell types either in direct cell-cell contact or at a distance as the hydrogel permits diffusion of soluble compounds. Notably, we show that co-culture of a breast cancer cell line with bone marrow stromal cells enhances 3D growth of the cancer cells in this system.
Highlights
Well-established methods to generate spheroids are spinner flasks, liquid overlays (LO) and the hanging drop (HD) technique.[8,9,10,11] the HD and LO techniques yield sized spheroids, but it remains a challenge to generate several thousand sized cell aggregates.[12,13] micro-structured surfaces with multiple microwells at the bottom of one single culture vessel represent an important alternative approach.[14]
We demonstrate that conical agarose microwell array (CAMA) is a versatile tool for 3D cell culture experiments examining growth measurement, treatment response and co-culture, and facilitates histological sample processing
Paper electron microscopy revealed the high accuracy in shape obtained by milling of the POM master structure and its subsequent moulding into PDMS (Fig. 1B). μCT scans of the final CAMA and of epoxy resin replicates confirm the precision of the replica moulding process (Fig. 1C, Video S1†)
Summary
Well-established methods to generate spheroids are spinner flasks, liquid overlays (LO) and the hanging drop (HD) technique.[8,9,10,11] the HD and LO techniques yield sized spheroids, but it remains a challenge to generate several thousand sized cell aggregates.[12,13] micro-structured surfaces with multiple microwells at the bottom of one single culture vessel represent an important alternative approach.[14] Agarose is a low-cost, transparent and nontoxic hydrogel, that is permeable to gas and small biomolecules. Flat bottom agarose microwells for 3D cell culture are useful for drug testing,[15,16] whereas pyramid-shaped agarose microwells allow stem cell aggregation and embryonic body formation.[17]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
