Unconventional technologies for plate loading aim to increase screening throughput and reduce wasted compounds and consumables
Hindered by the limitations of traditional approaches, companies are developing new technologies to increase the throughput and efficiency of screening experiments. Advancing to the forefront is acoustic drop ejection for liquid transfer.
Controlled drop ejection using sound has existed since the 1970s and has been investigated as an alternative to standard inkjet printing technologies, such as piezoelectric and thermal bubble. However, “no one was able to get that process to compete effectively with other inkjet technologies,” says Richard Ellson, chief technology officer with Labcyte Inc., Sunnyvale, Calif.
Moving tiny drops of ink from a reservoir to a sheet of paper is similar to moving tiny drops of a biological compound to a microplate well or other surface. Thus, printing technologies such as piezoelectric have been adapted for liquid handling applications in the life sciences. In 2000, Picoliter Inc., which has since merged with Labcyte, began investigating using acoustic technology to transfer biological materials.
The biggest advantage of acoustic is that it allows for the extraction of extremely small amounts of material, says Ellson. Labcyte’s current offering, the Echo 550 compound reformatter performs direct microplate-to-microplate transfers of droplets down to 2.5 nL (recently improved from 5 nL), “but the technology capability greatly exceeds that. . . . We’ve tested the technology down into the single picoliter range.”
Another advantage of acoustic technology over piezoelectric is that it does not touch the material being transferred. “In life sciences, you don’t usually know the physical properties of the material you’re working with . . . It is difficult to optimize conventional printing technology when the materials you are investigating have to come in contact with the device transferring them,” says Ellson. With acoustic, there is no contact between the compound and the transfer device, and the device does not need to be cleaned between compounds, thus saving on consumables.
Moving to uHTS
Tim Spicer, research scientist II, works as part of the core automation team in the lead discovery and profiling department at Bristol-Myers Squibb and is responsible for integrating new technologies into BMS’s automated platforms. Currently standardized around 384-well plates for its high-throughput screening experiments, the company is transitioning to ultra high-throughput screening (uHTS) with 1,536-well plates.
“Probably within a year, you’ll see us totally screening a 1,536-well format,” he says. Accomplishing this requires integrating three critical technologies into the company’s Thermo Electron Corp. CRS rail platforms, says Spicer. First is a robust compound reformatter, which here is the Echo 550. Second is a robust reagent dispenser, and at press time, BMS was contemplating the PerkinElmer FlexDrop or the Genetix AliQuot. Third is an imaging instrument; here, BMS was contemplating the PerkinElmer ViewLux or Leadseeker from Amersham Biosciences (now part of GE Healthcare).
BMS provided input and feedback to Labcyte during the development of the Echo 550. “We had telephone meetings with them every two weeks. … They had a set of criteria which were a challenge for us to meet. But we knew that when we met them, we would also be meeting [the needs of other companies] because BMS was probably a little tighter on the specifications than others are on HTS,” says Elaine Heron, PhD, CEO with Labcyte.
Ultimately, BMS served as a beta tester of the instrument, first examining it at Labcyte’s facilities in October of 2003. “We saw it dispense into 384- and 1,536-well plates, really just walking up. . . . It did very, very well. At that point, it dispensed with better precision and accuracy than any other instrument I’ve ever seen,” says BMS’s Spicer.
Waste and dead volume
Apart from not needing to be washed between compounds, Spicer points to another advantage of acoustic technologya reduction in the amount of wasted compound. “When you’re aspirating with a piezoelectric tip, typically you have to aspirate a microliter, dispense a few nanoliters, and then get rid of the rest. With the acoustic technology, you can dispense in 5 nL increments and save the rest [for later experiments].” BMS plans to upgrade its Echo 550 instruments to have a minimum drop size capacity of 2.5 nL in early 2005.
This is not to say that the technology eliminates dead volumes. “Their current standard [source] plate is a 384-well polypropylene flat-bottom plate that requires 20 µL in the well in order to be able to dispense properly,” says Spicer. BMS is working with Labcyte to develop plates that will reduce dead volumes. “We’re trying to develop a low-volume 384-well source plate, as well as 1,536-well source plate. I think a number of pharmas are racing toward 1,536-well storage.”
A byproduct of acoustic drop ejection technology enables users to ascertain the quality of their source plates. This is accomplished by examining the data the instrument generates when determining how much acoustic energy it needs to deliver and where on the liquid surface it needs to focus that energy to achieve a specific drop size. It is very difficult, not to mention impractical, to determine how much fluid is in a well simply by looking. However, by timing the reflection of sound waves off the surface and measuring the strength of the echoes, both the depth and the dimethyl sulfoxide (DMSO) content of the fluid in a well can be calculated.
Knowing the depth of liquid in a well can help avoid false results. “Some pharmaceutical companies confess that they occasionally run high-throughput screens where there wasn’t any material in the source plate. So they transferred nothing to the receiver plate and found that they had no hits. No interesting drugs came out because no interesting drugs went in,” says Ellson.
Furthermore, because DMSO absorbs water from the atmosphere, determining the DMSO concentration in a well can be difficult. “Someone might put together a plate where’s there is very little water. As they open the plates to transfer out of them, water from the atmosphere collects in the DMSO. So this is the first tool that really allows them to rapidly determine how much water has crept into their fluid,” he says.
The Echo 550 saves this auditing data in a spreadsheet format, says BMS’s Spicer. “It creates a file that tells you each well’s DMSO content, and it tells you if it did not [transfer] because a well didn’t meet the [minimum required] fluid thickness.” Labcyte plans a beta release of an instrument focused solely on auditing applications in the first quarter of 2005.
A twist on piezo
Palo Alto Research Center Inc. (PARC), Palo Alto, Calif., a subsidiary of Xerox Corp., is following another approach in advancing liquid handling technology. The company has developed a piezoelectric ejector, or “print head,” that integrates a fluid reservoir, piezo-actuator, and nozzle in one self-contained package.
“Commercial print heads have one reservoirone common fluidwhich goes to hundreds of nozzles. We have one reservoir which goes to one nozzle and a much smaller print head,” says John Fitch, mechanical engineer and area manager with PARC. The print heads double as a storage vial, holding up to 76 µL of fluid. Multiple print heads can be grouped together to load 96-well plates.
“We intend them to be disposable. You fill these things up with your precious fluid, and when it’s depleted you just toss it,” says Fitch. Fluid use is about 96%, and drop volumes are from 50 pL to 10 nL, with control of ± 2%, according to PARCs Web site (www.parc.com). The company is seeking partners to co-develop and customize this technology.
PARC is also secondarily investigating acoustic droplet dispensing technologies similar to Labcyte’s. “Our bio efforts have been mainly focused on the piezoelectric printing technology. The acoustic stuff works. We’ve done some work with it. But we have not refined that to a developed level,” says Fitch.
Labcyte’s Ellson, a founder of Picoliter, was previously an area manager in novel printing technologies at PARC.
Filed Under: Drug Discovery