TopSpot Vario: A novel microarrayer system for highly parallel picoliter dispensing

This thesis reports about the conceptional design, development and characterization of a novel system for production of microarrays, a so called microarrayer system. In the recent years microarray technology had a high impact in basic research, diagnostics and drug discovery. Its success caused tremendous activity of different companies to provide the most suitable microarrayer system. The here presented microarrayer bases on the commercially available TopSpot technology and is called TopSpot Vario. The TopSpot technology is an alternative to the piezo tip and pin printers which are currently dominating the microarrayer market. Its main advantage is the parallel ejection of up to 96 different samples and its one to one format change from the microtiterplate ordered reservoir pattern to the microarray spot pattern. The nozzles of the system are fabricated very homogeneously and very precisely in parallel using silicon micromachining. This is a clear advantage compared to other systems using single part productions like piezo tips and pins with a wider spread of important geometries. <br> <br>The novel developed TopSpot Vario system improves the standard TopSpot technology by replacing the pneumatic actuation principle by a direct liquid displacement method. This was realized by a piezoelectrically driven deformation of elastomer material into filled cavities and nozzles placed in their bottom. <br> <br>Core of the TopSpot Vario technology is a silicon micromachined printhead as it is for the standard TopSpot technology. Up to 24 different samples can be filled into the reservoirs which are ordered in a standard 384 well plate pattern. The one to one format change from the 6 by 4 ordered reservoir pattern with a pitch of 4.5 mm to the 500 µm pitched microarray grid is provided by microchannels connecting each reservoir with the corresponding nozzle. During actuation a parallel ejection of small droplets, one out of each nozzle, takes place. <br> <br>Different printhead designs and concepts were pursued. All components of the printhead like nozzles, displacement chambers, microchannels, etc. were separately optimized by different test-structures. For example, the optimized geometries of a via structure which connects microchannels in different levels improved the successful self-filling rate of the standardly used geometries from 62 % to 96 % for the applied test procedure. All optimized geometries were integrated into a finally used TopSpot Vario printhead design which was extensively and systematically characterized by using statistical planning of experiments. It provides an elastomer placement into the printhead before the printhead is filled. This enables a reliable sealing of all microchannels from each other during filling and actuation. Hence a reliable exclusion of crosscontamination between the samples is guaranteed. Key of success for this approach was the invention of a new microchannel structure for bubble-free priming of blind channels. <br> <br>The actuation principle of TopSpot Vario allows a tunable droplet volume of aqueous solutions from 250 pl up to 1600 pl. Indeed, one major conclusion of this work was the recommendation to print multiple small droplets into each other to achieve larger spot diameters instead of ejecting only one large droplet. This was due to the measured decrease of array homogeneity with increasing droplet volume. <br> <br>Compared to the standard TopSpot system, TopSpot Vario allows a much more flexible tunability of droplet volume and dynamics, a smaller minimum droplet volume (TopSpot Vario: 250 pl; standard TopSpot: ~700 pl), a higher maximum printing frequency (TopSpot Vario: 400 Hz; standard TopSpot: 30 Hz) and dispensing of higher viscous media (TopSpot Vario: eta < 11 mPa.s ; standard TopSpot: eta < 2.4 mPa.s). Further advantage compared to standard TopSpot is that not all nozzles of a TopSpot Vario printhead have to be used in parallel, so that even arrays with a pitch higher than 500 µm are possible. On the other hand, a pitch of 250 µm is possible on nearly all substrate surfaces due to the small minimum droplet volumes. Finally the reliable exclusion of cross-contamination during actuation is a clear advantage compared to standard TopSpot, where a blocked vent hole can result in a mixing of the samples in the actuation chamber. The finally used 24 channel printhead design can be easily enhanced to fabricate a 96 channel TopSpot Vario printhead which is already commercially available for the standard TopSpot system. <br> <br>The characterization of the TopSpot Vario system showed an intra nozzle reproducibility of droplet volume of 1.9 % and an inter nozzle reproducibility of all 24 droplet volumes in one array of 7.5 % at the recommended droplet volume of 270 pl. Reproducibility of average droplet volumes of one array from experiment to experiment was measured at 5 %. With these values the TopSpot Vario system can compete with all the other state-of-the-art microarrayers. <br> <br>Still to be optimized is the filling performance of the TopSpot Vario printheads. There, the main focus has to be set on minimization or even better an exclusion of the present risk of filling failures due to trapping of air bubbles in the displacement chambers. During 2040 performed fillings, 31 (1.5 %) displacement chambers with a trapped air bubble were observed. Due to the fact that even one fail filled displacement chamber results in the need of cleaning and re-filling of the entire 24 channel printhead, overall 11 % of the printhead fillings showed at least one of that filling malfunction. <br> <br>The current state of development of TopSpot Vario allows the use of the presented technology for a highly flexible microarray production. It is ready for beta testing where an occasional cleaning and re-filling of the printhead due to filling failures canbe tolerated.

Author dc:creator

• Steinert, Chris

Contributors dc:contributor

• Zengerle, Roland

• MST
• Drop-on-demand
• Microarrayer
• blasenfreie Befüllung
• Genomik
• MEMS
• microfluidics
• microarrays
• bubble-free priming

Repository record source_url

https://freidok.uni-freiburg.de/data/2308

OAI identifier oai:identifier

oai:freidok.uni-freiburg.de:2308