A fourth upgrade will be made to the Swedish Human Protein Atlas to coincide with the Human Proteome Organization’s Seventh Annual Meeting in Amsterdam in August, according to the program’s vice director.
In addition, the official has teamed up with an NCI researcher who has developed a special tissue arrayer for creating frozen tissue arrays that could become commercially available later this year.
Fredrik Ponten told BioArray News this week that the latest version of the atlas will include twice as many antibodies, and will include new search features that will allow users to look up different proteins and to view them in relationship to all human genes, and according to which class of proteins they belong.
“There will a doubling of antibodies — there will be 6,000 antibodies instead of 3,000 antibodies,” said Ponten. “The atlas will include all human genes, we will have protein information inside, and if you search for something, all human genes will be in the atlas.
“We have also added protein classes, so you can do more combined complex searches, like, ‘Show me all transcription factors on chromosome 17 which are expressed in liver cancer,’” he added. “So it will not just be based on the antibodies, but also all human genes and protein classes.”
The Human Protein Atlas is funded by the Knut and Alice Wallenberg Foundation, a Swedish organization that funds scientific research. The first version of the atlas, containing 718 antibodies, was released in 2005. By the time the third atlas upgrade debuted last year, it contained 2,800 antibodies. Upgrades are made annually, and this year’s will be the largest the project has made to date.
Antibodies that eventually make it into atlas upgrades are explored by combining the high-throughput generation of affinity-purified antibodies with protein profiling in a multitude of tissues or cell types assembled in tissue microarrays. The program is run by the Proteome Resource center, which is split between the Royal Institute of Technology in Stockholm and Uppsala University.
“Our goal is to have a first draft of the human proteome by 2014.”
According to the center’s website, the main objective of the project is to produce specific antibodies to human target proteins using a high-throughput method involving the cloning and expression of protein epitope signature tags. The antibodies are subsequently used to study expression profiles in cells and tissues, and for functional analysis of the corresponding proteins in a wide range of assay platforms.
“Our goal is to have a first draft of the human proteome by 2014,” Ponten said. Getting there, though, is going to take superior microarray tools. The atlas itself is not directly related to array-based research. It’s designed for “basic scientists working on protein biomarkers; cancer researchers in general”, Ponten said.
However, tissue microarrays have been integral to the creation of the atlas. The project collects material from surgical pathology both from normal tissues and cancer tissues, and constructs cell arrays from in vitro cultured cell lines.
The tissue and cell arrays are stained with all the antibodies produced by the project, scanned, and then annotated either by pathologists for tissue microarrays, or image analysis software for cell arrays. In 2006, BioArray News spoke with Caroline Kampf, site director at the project’s Uppsala-based facility, about the project’s use of tissue arrays (see BAN 5/16/2006).
The Swedish Human Protein Atlas uses three tissue arrayers sold by Beecher Instruments, an automatic ATA-27, a Manual Tissue Arrayer, and Beecher’s semi-automatic Galileo TMA CK3500. The project also uses Aperio scanners. Ponten said that the project’s current instrumentation is “sufficient,” but that it will take enhanced capabilities to reach the project’s objective.
“For me, since this is so high throughput, I would say that the bottleneck is getting good tissue from the beginning to use as donor blocks,” he said. Specifically, Ponten said that the project needs to be able to make arrays from frozen tissue. Typically, the project used tissue arrays comprising formalin-fixed, paraffin-embedded samples. However, over the past decade, more and more samples have been frozen with antigen retrieval rather than stored on FFPE blocks.
To deal with the bottleneck, the project has joined forces with Stephen Hewitt from the National Cancer Institute’s Tissue Microarray Research Program. Hewitt has developed a special tissue arrayer for creating frozen tissue arrays.
He told BioArray News in an e-mail this week that conventional approaches to making frozen tissue microarrays are “crude” and rely on either spotting the arrays adjacent to dry ice or in a cryostat. Hewitt had already developed an instrument called the “template arrayer” that spots tissues with a fixed template. To create the “cryoarrayer,” Hewitt needed to add temperature control features.
“Cryo is not trivial,” he said. “You need temperature controls for both donor and recipient. Not just cold, but controls. Cold needles are required as well,” Hewitt added.
According to Hewitt, patents for both the template arrayer and croarrayer are pending in the US. Hewitt developed the template arrayer together with Robert Starr, an investigator in the kidney disease section of the National Institute of Diabetes and Digestive and Kidney Diseases, and Lance Liotta, a professor of life sciences at George Mason University.
Now, the inventors are negotiating licensing and production deals for the template arrayer and cryoarrayer with Pathology Devices, a Westminister, Md.-based company that already sells a tissue microarraying instrument called the TMArrayer.
Hewitt said that he hoped that talks with Pathology Devices would facilitate the market debut of the systems this year. Ponten said that the Human Protein Atlas project is likely to begin using the cryoarrayer within a month or two.
“I was ignoring the cryoarray until [Ponten] convinced me he really needed it,” Hewitt said. “People talk about it, but really needing it is different. Frozen tissue is delicate, and histology is limited. But there are times where it is worth it.”
According to Ponten, the availability of the cryoarrayer will give the project new opportunities to characterize antibodies for the atlas. “Some antibodies don’t perform optimally in FFPE and proteins denature in FFPE,” he explained. “It will be nice to look at frozen tissue where you have more of a native protein. So I think it is a complement for those antibodies that are troublesome.”