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Open-source laser fabrication lowers costs for cancer research

26 January 2016

Researchers produce up to 50,000 microwells per hour by developing hardware and software modifications for a commercial CO2 laser cutter.

Rice's cost-saving microwell fabrication technique uses a commercial CO2 laser to fire millisecond laser pulses at a sheet of polydimethylsiloxane, a silicone-based organic and biocompatible polymer (photo: Jeff Fitlow/Rice University)

In a move that slashes 90 percent of the cost of mass-producing metastatic microtumours and therapeutic microtissues for screening and research, Rice University bioengineers have adapted techniques from the 'maker' movement to reprogram a commercial laser cutter to etch up to 50,000 tiny 'microwells' per hour into sheets of silicone.

The fabrication technique, which was developed with open-source software and hardware, is described in a new study published in the journal, RSC Advances.

"Microwells can be used to grow tiny clusters of cells," said Rice bioengineering researcher Jordan Miller, the lead researcher on the new study. "These clusters, or multicellular aggregates, contain 50-100 cells and have many potential applications, but they have been difficult to mass-produce. In the field of cancer biology, multicellular aggregates are used to model cancer micrometastases - small microtumours that can occur in high numbers in cancer patients. And in the field of regenerative medicine, they can be used as living building blocks to fabricate tissues and organs from a patient's own cells."

To study micrometastases in the lab, researchers grow multicellular aggregates of tumour cells. Traditionally, scientists have formed these by manually placing individual droplets of cells onto a plate using a pipette. But Miller said this method is labour-intensive, highly variable and typically produces small numbers of usable samples, which makes it impractical for studies that may require thousands of aggregates.

"Recent studies have revealed that cancer patients can have microtumours throughout their bodies," says Miller. "Most of these remain dormant, but some will actually grow into a full-blown tumour that can threaten the patient. We don't know exactly how the environmental conditions around a microtumour can promote or suppress this dangerous transition, but one way to investigate this process is with screening studies that involve large numbers of aggregates placed into defined environments."

One method for making many multicellular aggregates at once is to place cells onto a test plate containing several thousand microscopic wells, or microwells. Although commercial microwell products are on the market, Miller said they can be expensive, and the wells also come in a limited selection of shapes and sizes, which can make it difficult to produce small aggregates.

Miller and graduate student Jacob Albritton, the lead author of the new study, found they could produce up to 50,000 microwells per hour by developing hardware and software modifications for a commercial CO2 laser cutter - the same kind of machine used to make trophies, toys, acrylic figurines and other commercial products.

"We found we could create a conical well with a millisecond laser pulse in a sheet of polydimethylsiloxane [PDMS]," Albritton says. The material is a silicone-based organic polymer that's commonly used in industrial fabrication. This polymer is also non-toxic to living cells and is used often in biological experiments.

The researchers say their laser-based fabrication method can achieve something that standard lithography cannot: sharp, conical depressions that help guide the cells into a single aggregate. Moreover, it only costs around $30 to fabricate 100,000 microwells - less than one-tenth the cost of commercial sources.

The team found it could produce wells of different depths and shapes simply by varying the power and duration of the laser pulses, a technique that could produce specialised microwells that haven't been available previously.

"We theorised and then demonstrated that the focal plane controls the shape of the microwell that can be fabricated, and our new software modifications allow ultra-high-throughput production," says Miller. "We're using knowledge from electrical engineering to override the electronics of the laser cutter with open-source hardware and software.

"We bring in knowledge of biomaterials to etch and surface coat the silicone membranes, and there's also a good deal of cell biology and bioengineering that's needed to grow the multicellular aggregate tissues. Computer vision algorithms enabled us to automate analysis of tens of thousands of micro-tissues we can produce every day."

The team has open-sourced its hardware designs and software modifications by sharing them via Github, which empowers other groups around the world to reproduce the technique. While some biomedical labs might balk at purchasing and rewiring a commercial laser cutter, the team has also demonstrated a way to make polyurethane casts of completed sheets of microwells. Using these polyurethane microneedle templates, a lab could then make as many copies of microwell sheets as needed simply by casting new silicone over the microneedles.

"Our next studies here at Rice will investigate early steps of cancer progression as metastatic microtumours begin to infiltrate the surrounding tissue," Miller says. "And we've already distributed microwells to several other research groups around the country who have expressed interest in collaborating."

A YouTube video of the process is available to view here.


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