Microfluidics for 3D dynamic cell cultures in 384 multiwell format

Drug development is a lengthy and costly process, characterized by a high failure rate due to unreliable pre-clinical models, such as 2D cell cultures, which fail to replicate complex biological systems. 3D cell cultures and organ-on-a-chip technology offer more accurate representations of human tissues, replicating key factors like cell interactions and blood flow. Miniaturization and standardization are key for advancing these technologies. Combining 3D printing with Polydimethylsiloxane (PDMS) enables the creation of custom-designed prototypes that enhance the precision, scalability, and reproducibility of these advanced models. Here, we present a proof of concept of a well prototype, 384 multiwell format, constituted by an inlet connected to a syringe pump and an outlet, made of two components: an 8 mm 31 G needle and a 3D printed PLA dropper. The prototype was investigated by studying droplets volume, the relation between the ‘dripping rate’ and ‘inlet flow rate’ and the maximum sustainable flow rate. Three different fluids were used for these analyses: distilled water, phosphate buffer solution (PBS) and cell culture medium. The results showed that all three droppers exhibited similar behaviour across different fluids. At flow rates above 1 μL/min, the dripping time increased as the flow rate decreased, as expected. However, at lower flow rates, the dripping time was nearly twice as long due to flow inertia, causing a delay in droplet formation. The operational range of the droppers varied, with the 1.5 mm and 2 mm droppers showing a narrower range compared to the 2.5 mm dropper, especially when using cell culture medium. This suggested a correlation between the dropper’s performance and the type of fluid used.