February 15, 2006 (Vol. 26, No. 4)
Trend Toward Preplating and Preformatting
When you go to the grocery store these days, its all about convenience. Food is precooked and prepackaged into meals, so you can spend time doing other things. Perhaps then, it is no surprise that similar trends are occurring in life sciences support businesses.
A significant trend in the industry, says Charles Powell, vp marketing/sales at Aurora Discovery (www.auroradiscovery. com), is that suppliers will deliver content in a format thats more user friendly. That is, they will preload microtiter plates and ship them with material already loaded, whether it is kits, assays, libraries, or reagents. This approach can save time and reduce variability and error. It also requires less investment in automation equipment by the end-user.
As an example of this trend, Ambion (www.ambion.com) launched a new line of preplated, ready-to-transfect siRNA libraries in July 2005. Although Ambions target audience for the product was the segment of the market that doesnt have liquid-handling systems, pharmaceutical companies have also shown interest in the product because of its convenience, says Kathy Latham, Ph.D., senior marketing manager for RNAi and miRNA. The plates can be stored in the freezer or ordered on demand when screening is ready to begin.
According to Dr. Latham, an even bigger trend is Ambions custom formatting of its 1-nmol and 2-nmol siRNA libraries, a service that has been available for about a year. People love the idea that they can get the library plated exactly how they want, notes Dr. Latham, who points out that different customers have varied requests for plating formats.
Some are concerned about edge effects, some want wells left empty for controls, some want multiple aliquots, and others want to use 384-well instead of 96-well plates, she says. Although the siRNAs must still be diluted prior to transfection, avoiding the step of rearraying siRNAs for library screening is a huge benefit for the end-user, explains Dr. Latham.
Ambion has moved its production line to accommodate complete flexibility in arraying for its customers, Dr. Latham adds. The company has liquid-handling systems and LIMS to plate master stocks to customer specification and track everything.
GE Healthcare (www.gehealthcare.com) received positive feedback from market research before launching its Biotrak Easy ELISA technology in late 2004. According to GEs market survey directed toward scientists in basic research and lead profilers in drug discovery, anything that can enhance precision and save time is of value, says Rhona Macdonald, marketing director for lead discovery reagents.
Researchers can build the assays from scratch, but many choose to buy kits from assay providers. Microtiter plates that are precoated with an antibody of interest have been commercially available from several companies for a number of years. The researcher adds sample, detection antibody, and substrate to the wells with two wash steps required to perform the assay.
The new GE Healthcare Biotrak Easy ELISA technology is about making the process easier and saving hands-on time for the end-user, says Macdonald. In the new version, the microtiter plates include a prediluted standard curve and experimental wells that contain all the reagents necessary to conduct the assay. All the reagents are preformatted and lyophilized in 96-well plates. The researcher only needs to add sample and substrate to the wells. This new format results in time saving and increased precision, says Macdonald.
Another example of custom preformatted reagents is BD Biosciences (www.bdbiosciences.com) Lyoplate Technology, launched in September 2005. Lyophilization itself is not new, the key value is the quality and versatility of the proprietary matrix we are employing, notes Tony Ward, director of global strategic marketing for research cell analysis.
Custom microtiter plates that contain standardized and stable lyophilized reagents (Mabs, peptides, or cells) can be designed and engineered to order in just a few weeks. Benefits are simplified, ease-of-use, as well as potential reduction of error that can be associated with complex experimental designs.
Although the technology has broad applicability, one new example offered by BD is PhosFlow analysis, a flow cytometry-based technology employing phosphorylation-site specific antibodies and cell surface markers that allows activation state analysis of multiple proteins at single cell levels. Ward believes the technology has the potential to standardize clinical trials due to its ability to rapidly deliver customer specified kits.
Applied Biosystems (ABI; www.appliedbiosystems.com) offers customizable and preformatted 384-well microfluidic cards for real-time PCR. Customers can design their own TaqMan Low Density Arrays over the web by choosing from over 40,000 predesigned TaqMan Gene Expression Assays. ABI then manufactures the arrays with the chosen assays in the wells, explains Nico Tuason, product manager.
The microfluidic cards are used with ABIs 7900 HT real-time PCR system. According to Criss Walworth, product line director for gene expression, the TaqMan Arrays eliminate the need for liquid-handling robotics, since sample and reagents only need to be loaded into eight loading ports that each serve 48 wells. For studies done at multiple sites, the arrays offer additional advantages, according to Walworth, including highly reproducible and operator-independent results.
In December, the company expanded its offering of preselected, preformatted arrays to include the human G protein-coupled receptors panel, human protein kinase panel, mouse immune panel, human immune panel, and human endogenous controls panel.
Walworth says that ABI collaborated with pharmaceutical companies to choose the lists of targets that are included in the panels and that it was relatively easy to get agreement from the research community. Because the preformatted panels are offered as an inventory product, they can be ordered on demand. Another advantage is that they can eliminate the process of repeatedly designing and configuring assays.
People get hooked on convenience products, says Walworth. Sure they can do the work, but this makes it easier to do the actual experiment, notes Tuason.
Neal Cosby, Ph.D., manager at Promega (www.promega.com), agrees that people performing screens and assays want less hands-on time and more automation. One of the things they are saying is we want to save time in assay set-up, he notes. As an example, Dr. Cosby cites Promegas new Maxwell16 System for automated sample genomic DNA purification preparations from blood, tissue, mouse tails, or cells. Cartridges are prefilled with reagents. The Maxwell16 Instrument allows a hands-off approach, the researcher only needs to add sample and press Start. According to Dr. Cosby, Promega is currently looking into the idea of preformatting more assays in its portfolio.
Aurora Discoverys Powell believes that the above examples are evidence of a great opportunity in the market from a trend perspectiveto preload plates and ship them with material already loaded. The company is currently in discussions with kit and assay providers to develop pre-plating solutions that can temporally and spatially separate rare or special reagent management and assay set-up from assay performance.
As an example of this approach, many pharmaceutical companies currently perform semi-annual plating campaigns to create just-in-time libraries of screenable plates. As little as 10 nL of each compound is pre-dispensed into 3,456 well plates and stored for screening at a later date.
Aurora has expertise in piezo- and solenoid-valve microfluidic dispensing systems, microtiter plates, and detectors, according to Lane Niles, Ph.D., scientific fellow at Aurora. Auroras assay miniaturization technologies allow content to be accurately dispensed at small volumes (e.g., 10 L or under 100 nL).
The company is working to expand preplating applications. The idea is to preload the final work article with proper arrangement and array, says Powell. We think the next phase will bring more content onto the plate, eliminating other steps requiring exotic liquid-handling instrumentation by the end-user, he says. Greater value in R&D efforts will be realized by preformatting of the special materials into formats closest to or most ready for construction into assay samples, adds Dr. Niles.
Oleg Kornienko, Ph.D., senior research fellow in the department of automated biotechnology at Merck (www.merck.com), says that the company uses components of Auroras technology for compound management and uHTS. He points out that it takes some know-how to properly preplate and store small molecule compounds and aqueous-based reagents for later use.
People spend years refining the issues relating to pre-plating reagents so if that can be provided as a ready kit, thats huge, Dr. Kornienko comments. Sometimes biologists dont want to deal with instrument troubleshooting.
Other companies, such as Reaction Biology (RB; www.reactionbiology.com) and BioTrove (www.biotrove.com) leverage unique technologies to tackle the challenge of assay miniaturization.
Using the same printing technology that is utilized in constructing DNA microarrays, RB prints chemical compounds onto microarray chips in a volume of 1 nL, says Haiching Ma, Ph.D., CTO. With this DiscoveryDot technology we can put more than 6,000 chemical compounds on top of one 1×3 microarray chip to perform solution-phased biochemical functional assays, says Dr. Ma.
About four chips can fit on a microtiter plate, so about 25,000 compounds can be screened within the footprint of a microtiter plate. RB employs an aerosol deposition machine to activate the microarray-based reactions.
This technology has been validated for targets, such as serine proteases, cysteine proteases, metalloproteinases, histone deacetylases, HDAC, P450, phosphatases, and various kinases. Dr. Ma says that RBs technology is unique because the compounds and biological targets are always in solution.
The trick is to include 10% glycerol so that little tiny drops adhere to the arrays, he says. The arrays can be preprinted and frozen for later screening. RBs current model is to plate the libraries and conduct the screens for pharmaceutical companies as a service. But, they would also consider transferring the technology to a pharmaceutical partner.
BioTrove has yet another approach to assay miniaturization. The companys OpenArray application, based on Through-Hole technology, employs nanotiter plates with bottomless holes, each holding about 30 nL of liquid.
Any reaction you can do in a microtiter plate, you can do in an OpenArray, says Bob Pedersen, director of marketing, who points out that the array has all the benefits of solution-phase chemistry.
BioTrove launched OpenArray SNP genotyping analysis in September 2005, offering customers both preformatted and custom arrays. According to Pedersen, the platform has tremendous flexibility, and the company plans to expand into other areas of genomics, as well as the protein market.
The size of a microscope slide, each OpenArray contains 3,072 holes, which are divided into 48 different sectors that can be separately loaded by the user. Were trying to bring high-throughput to the masses by making it preformatted and easy to use without requiring costly robotics to achieve the same sort of density, says Pedersen.
Another benefit is significant cost savings. There is a 1,000-fold reduction in volume as compared to the same assays performed in 96-well plates. Although delivering 30 nL is not a trivial task, the OpenArray can be loaded without robotics, because the inside of the holes are hydrophilic and pull sample in. The hydrophobic rim of the holes keeps the liquid in place and prevents cross talk.