Biomedical Engineering Reference
In-Depth Information
Figure 1.5
Lysozyme crystals obtained on-chip at 5× and 20× magnifications.
1 µL of protein and 1 µL of precipitants were dispensed by programmed
Hamilton syringes [42] in each well of a 96-well plate containing paraffin oil.
Microbatch crystallization has been recently demonstrated in micropipettes
in 1 µL droplets by DeTitta's group at Hauptman Woodward Institute (HWI),
where the precipitant and the protein solutions are loaded manually into
a microcentrifuge tube, centrifuged, collected in a micropipette, and then
sealed [43]. Despite efforts to reduce the protein volumes, these processes
still consume a significant amount of protein and are labor-intensive.
Robotic automation has emerged as the dominant paradigm in state-of-the-
art high-throughput protein crystallization. However, robots are slow, very
expensive, and require high maintenance. Currently, there are only a few
automatic crystallization systems that are commercially available. Douglas
Instruments' Oryx 8 [44] can perform both microbatch and vapor diffusion
methods on protein samples in the range of 0.1-2 µL. Gilson's robotic work-
stations [45] can also perform both microbatch and vapor diffusion on protein
samples of about 1 µL. Syrrx, a rational drug design company, manufactures
a robotic system [46] for protein crystallization utilizing 20 nL to 1 µL protein
samples. State-of-the-art robotic systems at HWI's NIH-funded Center for
High-Throughput Crystallization have a throughput of 69,000 experiments
per day for setting up microbatch crystallization conditions, that is, a 96-well
plate could be setup every 2 min. Each screening condition still requires
0.4 µL of protein. These semiautomatic systems do not encompass ideal
high-throughput configurations, requiring user intervention for multiple
tray processing as well as suffering from other material-processing issues.
As most of the work performed with these systems is not on a large scale,
the automation of storage and handling of plates was not addressed in these
systems [47]. Such industrial systems, even though they are capable of setting
up thousands of crystallization screens a day, are prohibitively expensive for
academic research laboratories [48]. Therefore, affordable high-throughput
automation functionality of an industrial system is still needed.
Recent work has shown the feasibility of carrying out protein crystalli-
zation on digital microfluidic biochips. In [38], Srinivasan et al. presented
