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Immunology Center will continue to drive standard methods, better science

The Rochester Human Immunology Center (RHIC) has been awarded a $4 million renewal of its grant from the National Institute of Allergy and Infection Disease (NIAID). The renewal enables RHIC to continue leading the field of immunology in a worldwide effort to standardize how researchers use complex technologies like flow cytometry that are central to the discovery of new treatments.

When standardization is achieved, researchers will be better able to compare data collected worldwide, with the results of many studies combined into massive datasets to guide the construction of hyper-accurate computer models of the mechanisms of disease. Such simulations will yield scientific conclusions that are dramatically more valid and reproducible in areas like infectious disease, transplantation and cancer, researchers said. The resulting sophistication in understanding of the human immune system, and of how it responds to influenza, HIV or smallpox for instance, will guide the more precise design of near-future vaccines.

Since its founding four years ago, the RHIC has proven that it can drive the development of standard operating procedures, having helped more than 39 investigators and teams standardize their testing, and lending expertise that helped lead to the winning of 22 major grants. As an internationally known resource, the RHIC is part of working groups seeking to standardize methods for analyzing flu viruses and HIV in partnership with the Division of Microbiology and Infectious Diseases at NIAID, the HIV Vaccine Trials Network and the New York Influenza Center of Excellence.

"Standardizing test methods to achieve comparable results goes far beyond being able to compare apples to apples," said Sally Quataert, Ph.D., director of Core Facilities for the University of Rochester School of Medicine and Dentistry, and co-principle investigator for the grant renewal. "Validated methods assure researchers that their experimental results are good science, and truly meaningful. Standard methods also directly contribute to translational science, where the results of studies are more readily affirmed by regulatory bodies, and with fewer conflicting studies."

Prior to 2004, Quataert was director of Immunobiological Laboratory Services at Wyeth Vaccines Research in Rochester for seven years. Her industry background prepared her to apply Good Laboratory Practice (GLP) standards to academic studies, making them more likely to withstand scrutiny by the U.S. Food and Drug Administration as sound evidence that new treatments are safe and effective. If applied internationally, such standards would make drug candidates discovered at universities much easier to license to industry, with potential to increase the number of new drugs becoming available.

Also recently joining the RHIC executive committee was Tim Bushnell, Ph.D., director of the Flow Cytometry Core at the University of Rochester Medical Center. Bushnell's active role in the International Society of Analytical Cytology (ISAC) has positioned RHIC, not just to develop expertise, but also to take a leading role in the groups that are shaping related standards worldwide.

Flow cytometry is a method of counting and sorting cells that have been labeled by fluorescent markers which correspond with physical and chemical qualities of the cell. Modern flow cytometers can analyze thousands of cells in real time to unravel the complex interactions driving disease processes. The devices beam laser light into a stream of liquid which carries the cells to be analyzed. Detectors catch the patterns of fluorescent light that bounce off single cells as they pass through the laser. The cells have been prepared with fluorescent dyes that absorb the laser light and emit light at lower frequencies in scatter patterns that reveal information about the cell.

Modern instruments have multiple lasers and fluorescence detectors, with 18 fluorescence detectors now commonplace in larger research institutions. Increasing the number of lasers and detectors enables more precise identification of target cell populations by their characteristics. Four years ago, the RHIC had eight-color flow cytometry, but has since upgraded to 11- and 18-color cytometers with support from the Medical Center and its Clinical Translational Science Institute (CTSI). Researchers receive training on flow cytometry and other key discovery techniques and gain access to a growing suite of RHIC equipment. With the renewal of the NIAID grant, the center will continue to develop and standardize immunological methods.

Beyond general leadership and standard setting, the center will take the lead in the next five years in helping the field to make better use of three cutting-edge technologies "of immense importance." The goal is to advance technologies to maturity so that they can be applied widely, with the new standards communicated through publications, the RHIC Web site and symposia.

The first technology is arrayed image reflectometry, which promises to be very useful in large-scale analysis of immune responses against multiple disease-causing proteins like those encountered during influenza outbreaks. Immune system proteins can be coated with a thin film that prevents light from reflecting off of them, but that become more reflective as the proteins bind to disease-related proteins. When several proteins to be tested are attached to a biochip, a great many proteins can be analyzed for little cost.

The grant renewal will also bring together RHIC experts in cytometry and in a second new technology: quantum dot nanostructures. These microscopic machines have optical qualities that can be harnessed to measure cellular qualities via flow cytometry with more precision than fluorescent dyes. Like dyes, the dots can be designed to inhabit certain regions of the cell, or cling to certain proteins, so that their number and characteristics can be measured. Dyes are an older technology, limited in number and with overlapping emission spectra, which means they interfere with each other. RHIC teams will seek to establish new Q-dot production methods to replace and extend dye capability. As RHIC machinery gets more complex, it will become increasingly able to pick apart and to build accurate immune cell models.

Thirdly, the team will seek to design better "gating" procedures. Flow cytometry separates cell types by brightness into groups, and gating is the process by which researchers select which cells go into each group. The current process in many labs is arbitrary, tedious and has poor reproducibility between operators because research teams select patterns of cell characteristics "by eye." The field of flow cytometry is in desperate need of high-speed automation to process the "mountains of data" generated by the tests, and when gating the results, researchers said. The current process is greatly slowing the pace of discovery as research teams do double duty with lab work and time-consuming computational work.

One unique capability that drove the RHIC renewal was that the Medical Center has a world-leading core of biostatisticians along with leading immunologists. Led by Hulin Wu, Ph.D., chief of the Department of Biostatistics and Computational Biology, the biostatistics team within RHIC will seek to establish statistically-rigorous, automated gating protocols for flow cytometry analysis that will greatly increase the reproducibility of data and the speed of flow cytometry lab by lab.

"The NIAID reviewers that voted to renew the RHIC grant seem to have validated the fundamental concept of our center that centralizing the development and deployment of sophisticated technologies increases the efficiency of many other studies," said Tim Mosmann, Ph.D., principal investigator of the RHIC grant, and director and Michael and Angela Pichichero Director's Endowed Chair of the David H. Smith Center for Vaccine Biology and Immunology at the Medical Center. "Although research core facilities can provide centralized expertise on a fee-for-service basis, the RHIC grant allows us to move beyond this model by actively pursuing new technologies and developing promising methods via direct funding, and by providing expertise in human immunology on a collaborative basis. Secondly, the RHIC has a strong training function, so that expertise developed in the RHIC spreads rapidly through the research community. We are delighted that many researchers see the RHIC as an asset, and it is their achievements, publications and grant awards that were a major reason for the RHIC grant renewal."


Contact: Greg Williams
University of Rochester Medical Center

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