Monitoring for Aggregates
The issue of precipitates or aggregates in solutions is one that vexes nearly every lab. The Plate Auditor, cameras to look for precipitates in samples, and devices to measure the amount of water in a DMSO solvent, are some solutions that researchers use.
Robert Damoiseaux, scientific director of the Molecular Screening Shared Resource at UCLA’s Jonsson Comprehensive Cancer Center, uses dynamic light scattering to look for particles or aggregates in solutions using 384 and 1,536 well plates. Wells containing particles are flagged for further inspection.
“We use this for nanoformulations quite a bit,” said Damoiseaux. “For example, the compound Abraxane® is nanoparticles made of Taxol particles with an albumin shell. It’s a great concept, because Taxol is very hydrophobic and precipitates out really easily.”
The albumin shell renders it less hydrophobic, making it more potent and easier to infuse into patients.
“The vasculature around cancers actually actively recruits albumin because it’s bound to many nutrients,” explained Damoiseaux. “So basically the tumor loads itself up with the albumen-coated nanoparticles by the active transport mechanism, resulting in enrichment of Taxol where it needs to be.”
In compound management, nanoparticles are more needy than their chemical cousins. They are less stable than small molecules, which can exist happily in DMSO in a desiccator.
“But with nanoparticles, even if they’re in powder, they clump, they aggregate, they re-precipitate, there are templating effect. It’s a very complicated picture,” said Damoiseaux. “That’s something that dynamic light scattering is very useful for.”
Challenge of Dry Compounds
Recent progress in compound management has not, however, included the transfer of dry compounds, despite the high demand for automation of this common task.
“The physical transfer of dry compound from source to destination is still a very manual process, mainly because the industry has not developed an acceptable automated solution due to the extreme complexities of transferring dry compounds,” says Genentech’s Hascall.
“Although weighing compounds is one of the least desired tasks in the lab, it is a critical first step to preparing the sample for testing. Whether compound management is weighing a large library, or newly synthesized compounds from chemistry, it has historically required human manipulation of the compound for the transfer to occur.”
One of the challenges of automating this process is the type of solid—powders are much easier for robots to work with. Hascall’s group uses a fully automated weighing system called the Nova, from Innovate Engineering, to transfer dry, free-flowing powders.
Other solids such as dry films, oils, globular material, or chunks of crystals, still present challenges. For powder transfer, the Nova uses a stainless steel collector pin charged with static electricity, to which the powder clings.
The Nova changes the collector pin each time to avoid cross contamination, and uses powder level detection (similar to liquid level sensing) to calculate the static charge, and the right level at which to place the collector into the vial.
“Fully automated weighing holds many challenges that the Nova has been designed to handle,” says Hascall. “With a greater than 85% success rate in our study of over 4,000 compounds, the Nova can be considered to be one of the first fully automated compound transfer robots.”
It is also integrated into Titian’s Mosaic compound-management software to eliminate manual data entry.
Even though compound management is evolving quickly, new challenges are directly ahead. For example, nucleic acid reagents such as plasmids, siRNA, microRNA, and shRNA are increasingly stored and require a special kind of management.
A repository that stores plasmids, which are typically purified using RNases, and also stores siRNA must have separate, dedicated liquid-handling systems to avoid destruction of the siRNA.
“These are the strategies,” says Damoiseaux. “You have to be very careful to set up your equipment in such a way that the workflows are parallel, and the overlaps are such that you are not damaging reagents.”
Whether for nucleic acid reagents or other compounds, no doubt parallel advances in software and automated instrumentation, such as MScreen, the Plate Auditor, and the Nova, will pave the way to progress, with labs running more smoothly than ever.