Microstamped Petri Dishes for Scanning Electrochemical Microscopy Analysis of Arrays of Microtissues

Abstract

While scanning electrochemical microscopy (SECM) is a powerful technique for non-invasive analysis of cells, SECM-based assays remain scarce and have been mainly limited so far to single cells, which is mostly due to the absence of suitable platform for experimentation on 3D cellular aggregates or microtissues. Here, we report stamping of a Petri dish with a microwell array for large-scale production of microtissues followed by their in situ analysis using SECM. The platform is realized by hot embossing arrays of microwells (200 μm depth; 400 μm diameter) in commercially available Petri dishes, using a PDMS stamp. Microtissues form spontaneously in the microwells, which is demonstrated here using various cell lines (e.g., HeLa, C2C12, HepG2 and MCF-7). Next, the respiratory activity of live HeLa microtissues is assessed by monitoring the oxygen reduction current in constant height mode and at various distances above the platform surface. Typically, at a 40 μm distance from the microtissue, a 30% decrease in the oxygen reduction current is measured, while above 250 μm, no influence of the presence of the microtissues is detected. After exposure to a model drug (50% ethanol), no such changes in oxygen concentration are found at any height in solution, which reflects that microtissues are not viable anymore. This is furthermore confirmed using conventional live/dead fluorescent stains. This live/dead assay demonstrates the capability of the proposed approach combining SECM and microtissue arrays formed in a stamped Petri dish for conducting cellular assays in a non-invasive way on 3D cellular models.

Non‐Invasive Monitoring of Osteogenic Differentiation on Microtissue Arrays under Physiological Conditions Using Scanning Electrochemical Microscopy

“…Control microtissues are not supplemented with BMP‐2. Microtissues exhibit a stable size of 206±34 µm after 24 h in culture, as previously reported by us 21, and do not show significant size changes upon exposure to BMP‐2. Furthermore, individual microtissues can easily be retrieved from the platform using a micropipette.…”

“…The microwells are stamped in commercially available Petri dishes using a custom‐made set‐up and a PDMS mold. The stamped dishes are suitable for simultaneous SECM and optical microscopy 21. The stamped dishes are coated overnight with 1 % w/v (1 g in 100 mL) Pluronic F127 solution.…”

Isolated Reporter Bacteria in Supramolecular Hydrogel Microwell Arrays

“…Standard microfabrication techniques, including photolithography, 12 micromolding, 13 embossing, 14 and colloidal lithography 15 have been used for microwell fabrication. For instance, polyethylene glycol (PEG)-based microwell arrays fabricated using micromolding in capillaries, 13 polystyrene microwell arrays obtained by hot embossing, 16 or poly(vinyl alcohol) (PVA) microwell arrays prepared using photolithography 17 are illustrative examples. Microwell arrays can be adapted to facilitate the acquisition of detailed information about cell–cell and cell–environment interactions.…”

Electrochemical imaging of cells and tissues

“…The 3D cell aggregates of live HeLa cells inside microwells were analyzed using RC-SECM for O 2 consumption as a non-invasive live/dead electrochemical assay. 64 Oxygen concentration gradients inside and outside of a live brain slice, excised from mice, were monitored with platinized nanoelectrodes. 11 …”