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While deposition with the chip proved precise, switching to a dedicated line for cell delivery

put much less stress on the microfluidic device. Given its lengthy fabrication process,
preserving its functionality was a clear benefit. Furthermore, the dedicated line allowed a

higher fluid pressure and thereby a higher dispensing throughput (Error! Reference source
not found.). Given that liquid dispensing was required twice per culture well – once for pre-

filling and once for cell delivery –, the speed of liquid delivery was a major factor in liquid

dispensing.
Moreover, using the microfluidic device itself for dispensing would have resulted in a wasted

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volume of 20 mL of medium containing 4x10 cells for a 384-well plate, as cell-laden medium
would constantly flow to the waste channel to keep a constant flow rate while the dispensing

step is in progress. For the 1536-well plates, however, cell delivery via a dedicated line proved

challenging. First, the well dimensions were 1.7 x 1.7 mm², requiring the delivery line with an
outer diameter of 1.6 mm to be perfectly centered. While this was alleviated by adding a

dispensing tip to the end of the tubing, dislodging the droplet from the needle proved more
problematic, as the working volume of a 1536-well plate was specified with 4 – 12 µL. Initially,

the needle was kept so close to the well-plate that moving to the next well wiped the tip.
However, as the tip was not always positioned at the same height, it would get entangled when

moving the well plate, resulting in double dispensing and overflowing wells. In addition to

overflowing wells, wiping the nozzle between wells increased the risk of infection. For the
1536-well plates, cell deposition using the microfluidic device was therefore favored, as having

an air-pressurized outlet allowed to precisely eject even small volumes down to 10 µL
repeatedly.

Excluding drying and UV sterilization, the automated workflow for preparing a fully seeded

plate required approximately 20–25 min for 96 wells, 1 h 15 min for 384 wells, and about 5 h
for 1536 wells. However, manual steps still added substantial time to the overall process,

particularly the drying step, which took several hours. This step could be significantly
shortened in the future by replacing heat evaporation with direct liquid removal or vacuum-

assisted drying. Additionally, parallelization of plate handling or the use of alternative carrier
liquids such as alcohols, which evaporate faster and maintain sterile conditions, could have

further reduced preparation time. Of note, microscaffolds could have been presorted and stored

in the liquid, allowing the cell culture workflow to start when needed. Importantly, the design
of the device enabled a clear path toward scalability, as it allows to multiplex multiple units in

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