Improving care is a mission that drives us all. Translating new discoveries into better clinical outcomes takes dedication and new approaches to asking the most difficult questions. At Fluidigm, we believe that changing the course of how diseases are treated and ultimately cured requires a comprehensive understanding of complex cellular phenotypes and their interrelationships within the tissue microenvironment. To meet this great challenge, we developed the Hyperion™ Imaging System.

“The impact of this first-in-class technology stands to be especially profound in oncology, immunology and immunotherapy research. With more than 30 systems enabled globally, progress in applying this innovative technology to ask new questions across a number of disease research areas is truly impressive .”
—Chris Linthwaite, President and CEO, Fluidigm

The Hyperion Imaging System brings imaging capability together with proven CyTOF® technology to set a new standard in highly multiplexed protein detection. Surpassing the inherent limitations of fluorescence detection, this system enables researchers to simultaneously image 4 to 37 protein markers in tissues and tumors by Imaging Mass Cytometry™. Ideal for deep profiling of precious formalin-fixed, paraffin-embedded (FFPE) and fresh frozen tissue samples, the Hyperion Imaging System empowers researchers to uncover new biomarker correlations and cellular social networks in the context of the tissue microenvironment—all from a single slide.

As with any new research application, having access to optimized reagents, analytical tools and training is essential for success. We designed the Hyperion Imaging System workflow with this in mind. It is accompanied by a pathologist-verified Maxpar® imaging antibody portfolio, antibody labeling kits, a software suite for image acquisition and data analysis, and experienced Fluidigm service and support. With the ability to utilize up to 135 channels to detect additional parameters, the system is well-suited to meet researchers needs today and well into the future.

A step jump of this magnitude doesn’t happen every day in life sciences. The impact of this first-in-class technology stands to be especially profound in oncology, immunology and immunotherapy research. With more than 30 systems enabled globally, truly impressive progress is being made in applying this innovative technology to ask new questions across a number of disease research areas.

I invite you to join with us and the growing scientific community in powering this revolution in tissue imaging to create breakthroughs today that will enable the development of better diagnostics and more effective therapies for tomorrow.

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The Hyperion™ Imaging System received first place in The Analytical Scientist Innovation Awards, or TASIAs. Judges named it the top innovation of 2017, commending the platform that “combines tissue imaging with cytometry, allowing correlation between biomarkers and cell interactions. This technology has the potential to be truly transformative.”

Editor Charlotte Barker of The Analytical Scientist said, “Now in year five, The Innovation Awards continue to shine a light on the instruments and technologies that are having a big impact across the analytical sciences. We had a record number of entries this year and the final 15 reflect the full spectrum of analytical advances, from ingenious software for managing impurity data, to transformative MS imaging technology.”

The Hyperion Imaging System has the potential to revolutionize disease research by providing unprecedented visualization of complex cellular phenotypes and their relationships in the context of cancer, immuno-oncology and a range of immune mediated diseases. The potential ability to develop better diagnostics and more effective therapies stands to change the future of health care.

Read the full article in The Analytical Scientist for more on the TASIAs. To learn more about Imaging Mass Cytometry™ and be informed about news and events, fill out the form below.

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Take a moment to remember your favorite painting, a classic that brings you comfort and fills you with introspective thought. Something like Starry Night by van Gogh or Guernica by Picasso. Now imagine that instead of showing the full image, we break it all down inch by inch, pixel by pixel into a collection of dots grouped by color. Does knowing that 80% of Starry Night is blue or that 10% of Guernica is white convey the same message as seeing the image as the artist intended? Even though all the colors are represented, the work of art disappears in the absence of spatial organization. For David L. Rimm, MD, PhD, Professor of Pathology and of Medicine at the Yale School of Medicine, spatial information brings everything together for a better understanding of the cellular environment. That is the power of imaging in scientific studies. The new Hyperion™ Imaging System brings together imaging with proven CyTOF^® technology, pushing highly multiplexed immunohistochemistry-based imaging to the forefront of translational research. Rimm and colleagues are leading the way.

Recognizing early potential

Rimm’s familiarity with highly multiplexed imaging technology began in 2014 when he was writing a commentary on two groundbreaking Nature papers on novel imaging methods. Based on two different techniques, the publications coalesced on the same theme: the need for higher-order imaging technology using metal-tagged antibodies. Rimm recalled that the techniques “were similar approaches but they had some subtle differences in the mechanism.” Rimm recognized the value of these techniques and saw this as the future path for his lab. He saw great potential in the Hyperion Imaging System, where a precisely directed laser beam collects biological samples stained with metal-tagged antibodies and CyTOF technology enables the detection of different metal tags based on mass rather than on wavelength, which is the basis of relatively low-plex immunofluorescence.

For Rimm, this highly multiplexed imaging technology, Imaging Mass Cytometry™, represents the future of pathology-based studies. “A key current clinical problem in pathology,” said Rimm, “is the ability to define levels of protein in tissue so that we are maintaining the spatial information and the localization of different cell types.”

He added, “Research in my lab is related to biomarkers that either predict outcome or response to therapy or to a drug. I see the Hyperion Imaging System as a valuable discovery tool that can provide information about which molecules are most important to predict response to therapy. It gives you information that few other tools can provide: the ability to look at 30 different things at once yet maintain the two dimensional spatial information, giving us the power to discriminate biology beyond a single point measurement.”

Developing biomarker signatures

Rimm’s lab focused on applying Imaging Mass Cytometry technology to two innovative initiatives. The first was discerning key signaling pathways regulating biological effects of targeted therapies in breast cancer. Using banked samples from cancer patients treated with trastuzumab (Herceptin®), Rimm’s group simultaneously analyzed the expression levels of 17 protein markers using Imaging Mass Cytometry. By concurrently staining with a multitude of markers, he said, we can examine “the signaling molecules themselves, the receptors and the molecules in the signaling cascade that occur after the receptors are bound.”

For the second effort, they used Imaging Mass Cytometry to better understand the tumor microenvironment of lung cancer, with a particular emphasis on tumor infiltrating immune cells. This project used Imaging Mass Cytometry to develop a protein-based signature associated with patient response to therapy, with an ultimate goal to more precisely tailor therapy for cancer patients receiving immunotherapy.

The Hyperion Imaging System requires relatively small quantities of tissue to detect large numbers of proteins while maintaining spatial context. This is a key benefit when analyzing small biopsies or relatively rare or precious specimens. Rimm’s training set of tissues for the Herceptin study consisted of tumors from more than 61 patients.

Examining 17 markers simultaneously offered the advantage of identifying a wide variety of cells and activation states without the need for serial sections. Speaking to this point, Rimm said that when using serial sections, it’s unclear if the markers are actually in the same cell. It’s then necessary to take a population approach comparing expression of markers 100 µm apart. Imaging Mass Cytometry on the Hyperion Imaging System provides “unequivocal evidence that they are in the same cell, because they co-localize to the same pixel.”

“I see the Hyperion Imaging System as a valuable discovery tool that can provide information about which molecules are most important to predict response to therapy. It gives you information that few other tools can provide.”
—David Rimm, MD, PhD, Yale

Looking ahead

For Rimm, the real power of this technique will come from uncovering unanticipated correlations between large numbers of protein markers and patient outcome. Imaging Mass Cytometry enables simultaneous analysis of large numbers of protein markers to more efficiently discover new predictive signatures composed of smaller numbers of markers that may ultimately have clinical utility.

Scientists can also use Imaging Mass Cytometry to identify relevant biomarkers of disease progression and to better understand how cancer arises. When faced with the complexities of a heterogeneous disease state, looking at as many markers as possible can create insightful results that could ultimately lead to improved prognosis and therapies—and by extension improved care for cancer patients.

Going forward, Rimm hopes that Imaging Mass Cytometry will “enable us to answer key biological questions.” Moving away from the one-dimensional data of mass cytometry experiments into two or three-dimensional spatial data will advance future biological exploration. With a sense of reserved excitement, Rimm said he believes that “we’re just at the very beginning of finding out what the Hyperion Imaging System can do.”

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The innovative Hyperion™ Imaging System by Fluidigm was born out of a dedicated effort to advance translational research toward a more comprehensive understanding of the complex cellular relationships and tissue microenvironment. It is the first commercially available platform to empower simultaneous interrogation of 4 to 37 protein markers in spatial context at subcellular resolution. A lead developer and longtime champion of Imaging Mass Cytometry™, Bernd Bodenmiller, PhD, Assistant Professor for Quantitative Biology at the University of Zurich (UZH), shares his experiences using the Hyperion Imaging System.

In a Nature Methods paper published in 2014, Giesen and Wang et al. first reported on an approach they developed in the Bodenmiller Lab to simultaneously image 32 protein markers and protein modifications at subcellular resolution. They detailed an Imaging Mass Cytometry approach they believed would greatly advance the understanding of cancer and other disease states through improved tissue characterization. Imaging Mass Cytometry now brings game-changing technology to immuno-oncology researchers, with the realm of possibilities extending to autoimmune diseases, neuroscience, precision medicine and other disease areas.

At his systems biomedicine lab at the north end of Lake Zurich in Switzerland’s largest city, Bodenmiller sets his sights on understanding the workings of tumor ecosystems. The Bodenmiller Lab researches cell types present in solid tumors to better grasp how the cellular elements of the ecosystems influence each other through cell-to-cell communication to drive tumor development. Modeling the tumors as multidimensionally resolved cellular networks, Bodenmiller studies cancer biology in the context of precision medicine, hoping to inform future treatment approaches for cancer. His comprehensive investigational research encompasses development of experimental and bioinformatics methods for highly multiplexed, spatially resolved subcellular tissue analysis.

The Imaging Mass Cytometry workflow pairs immunohistochemical methods with proven CyTOF® technology to record cellular markers, transcripts and signaling to generate an image of the tissue. Having already published a paper demonstrating how Imaging Mass Cytometry enables the simultaneous imaging of 32 protein markers at subcellular resolution in breast cancer tissues, the team has also shown the simultaneous imaging of proteins and transcripts, expanding the possibilities for analyzing aspects of cancer biology.

Images generated using the Hyperion Imaging System could reveal how cells respond to oxygen levels, factors contributing to immune cell exhaustion, and which conditions within the tumor environment relate to clinical features. Put into the context of translational research, the Imaging Mass Cytometry workflow using the Hyperion Imaging System empowers scientists to gain comprehensive insights into solving questions relevant to clinical treatment and global health initiatives underway at the University of Zurich.

Analyzing the tumor ecosystem

While all of his group’s research is based on mass cytometry, Bodenmiller estimates that around two-thirds of their projects now rely on Imaging Mass Cytometry. “The Hyperion Imaging System is a key method in my lab,” he said. “Within the tumor ecosystem it is informative to reveal cell type identity and the functional state of individual cells. It is important to also add spatial information for those cells. This is highly relevant because then we can study how cellular environments might actually shape a cellular phenotype or influence treatment response.”

Imaging Mass Cytometry can provide translational data on the status of the immune repertoire within a tumor. Bodenmiller anticipates that targeted modulation of the tumor ecosystem will eventually empower researchers to interrupt and halt tumor growth. “Immune checkpoint blockade therapies are a nice example of targeting interactions within the tumor ecosystem,” he explained, “as it’s really the cell-to-cell interaction and modulation that are responsible for dampening and reactivating the immune response.”

Comparing the way Imaging Mass Cytometry uses metal isotopes as reporters to the way enzymatic immunohistochemistry uses enzymes to stain tissue, and immunofluorescent microscopy uses fluorophore as a reporter for antibodies, Bodenmiller said, “What makes Imaging Mass Cytometry special is that it enables us to perform multiplexed measurements.” When detecting isotopes on the Hyperion Imaging System, the proven CyTOF technology offers 135 channels to measure many metal-tagged antibodies at once in a single tissue.

“I see enormous potential for Imaging Mass Cytometry in assessing immune checkpoint inhibitor status and dereguated cellular pathways and determining best treatments for precision medicine.”
—Bernd Bodenmiller, PhD, University of Zurich

Unprecedented pathology possibilities

Imaging Mass Cytometry stands to benefit many different areas of medicine, health and life sciences. With a nod to translational and potential clinical applications, Bodenmiller pointed to Switzerland and countries with centralized state-run health care systems that store anonymous clinical and long-term patient data and tissue samples. He cited Zurich as an example housing a half-million tissue specimens that scientists could use for additional research under governance by the institutional and state ethics board.

“There are untold answers in the millions of biobank tumor samples stored in hospitals around the world—many with densely annotated disease and patient features or treatment data,” he remarked, adding that there is an unprecedented amount of information to explore from tissue samples. “If we analyze large cohorts using Imaging Mass Cytometry and apply computational tools and machine learning to improve patient classification, we could learn about marker relationships that translate to established clinical techniques in the future. The potential is enormous.”

Toward precision medicine

Knowledge about tumor biology could be translated to patients in the future, Bodenmiller observed. “I see enormous potential for Imaging Mass Cytometry in assessing immune checkpoint inhibitor status and deregulated cellular pathways and determining best treatments for precision medicine. With Imaging Mass Cytometry, we gain an unprecedented detailed view of tissue biology. We have the ability to determine T cell status and which markers they express, suggesting how to modulate them within the tumor ecosystem. Our ability to comprehensively measure tumor tissues—from single-cell pathways to cell-to-cell interactions and tissue morphology—will reveal which aspects of the tumor ecosystem to target.”

Bodenmiller envisions the development of applications on the Hyperion Imaging System that could potentially optimize clinical workflows. A comprehensive understanding of tissue biology stands to benefit the development of new treatments and decisions about the right drug for the right patient. He contends that such precision medicine is close at hand. Using an example of analyzing melanoma samples with highly activated signaling pathways, he noted that it would be possible to target those pathways with appropriate inhibitors.

Nature Methods on histoCAT

In September 2017, the Bodenmiller Lab’s Denis Schapiro, Hartland Jackson and the UZH team published a Nature Methods paper describing the computational toolbox they call histoCAT™ (histology topography cytometry analysis toolbox) and documenting the processing of Imaging Mass Cytometry data to analyze cellular phenotypes and spatial relationships of these cells with neighboring ones. Fluidigm recently announced a distribution agreement with UZH to offer histoCAT software for high-parameter tissue analysis.

Bodenmiller expects other publications using histoCAT software analysis to follow: “We plan to show how to relate these Imaging Mass Cytometry results to clinical data, and in an additional extension of histoCAT, how we can analyze three-dimensional representation of tumors.” Advancing from two-dimensional tissue representations to reconstructing three-dimensional tumor volumes will provide an even more comprehensive understanding of cell-to-cell interactions.

Challenges and successes

There are many obstacles ahead in immuno-oncology research. We can answer more questions faster with Imaging Mass Cytometry to achieve our common goals for better human health, but developing this remarkable technology calls for the power of committed and engaged researchers focusing together on that shared objective. As with other big data, data-driven medicine analyzes massive sets of medical records and images for patterns that can reveal previously hidden connections to advance learning and inform researchers’ decisions. The answers are out there, and the tissue samples are out there, Bodenmiller concludes.

Bodenmiller reiterated the value of Imaging Mass Cytometry using the powerful Hyperion Imaging System and the potential it holds for researchers studying human health and disease. “My lab is part of the Cancer Research UK (CRUK) Grand Challenge. Imaging Mass Cytometry is one of the main pillars of this whole project with regard to comprehensively measuring the tumor ecosystem,” he said. “It is a Grand Challenge because the grant aims to comprehensively map a high number of tumors on a molecular and cellular level in 3D. About 10 labs in the US, UK and Switzerland are involved, and we’re attempting to build the whole pipeline and data analysis to enable this.”

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Tumor gene expression has proven effective in measuring immune response during cancer progression and therapeutic response. As emerging therapies reveal new biomarkers and expand the need for samples, the costs and labor required to complete this important work also rise. Cancer researchers now have a reliable, sensitive and cost-effective tool for identifying gene expression signatures from immune and cancer cells: the new Advanta™ Immuno-Oncology Gene Expression Assay, used with the Biomark™ HD system.

A powerful immuno-oncology focused assay

The Advanta Immuno-Oncology Gene Expression Assay was developed in collaboration with leading academic and biopharma researchers to provide the right balance of biomarker breadth, assay flexibility and workflow efficiency.

The panel consists of 170 genes, including 91 reported in Nature, where Roy Herbst et al. defined a gene set representing tumor and immune response. The panel includes markers for:

• Immune cell identification
• Immune and cancer cell function
• Immune regulation and cell fate
• Checkpoint therapy response

Optimized for FFPE and fresh frozen tumor samples, the Advanta Immuno-Oncology Gene Expression Assay uses TaqMan® chemistry to sensitively measure gene expression, with five reference genes serving as analysis controls. Combined with Fluidigm microfluidics technology, the assay uniquely offers significant workflow efficiencies over traditional gene expression profiling methods. Each reaction is miniaturized to nanoliter volume and controlled using precise automation to empower accurate and cost-effective qPCR analysis across a large dynamic range. Since assay introduction into the integrated fluidic circuit (IFC) is under user control, researchers also have the flexibility to add up to 17 new assays or exchange gene assays within the panel to achieve experimental goals—all without affecting the original panel content, protocol or workflow.

Immuno-Oncology Expression Assay

Table 1. The Advanta Immuno-Oncology Gene Expression Assay panels A and B. This two-panel set includes 91 target genes and 5 housekeeping genes on Panel A and 74 target genes and the same 5 housekeeping genes on Panel B. Panel B reserves 17 open assay inlets, so you have the added flexiblity to add new assays over time for customization. Each panel is available in packs of 2 or 10 (192 or 960 samples, respectively)

Sample case study

In a 2016 paper published by The Lancet for the POPLAR study, “Atezolizumab versus docetaxel for patients with previously treated non-small-cell lung cancer,” Louis Fehrenbacher et al. detailed how they “assessed efficacy and safety of atezolizumab versus docetaxel in previously treated NSCLC, analyzed by PD-L1 expression levels on tumor cells and tumor-infiltrating immune cells and in the intention-to-treat population.” The team showed that “patients with pre-existing immunity, defined by high T-effector–interferon-γ-associated gene expression, had improved overall survival with atezolizumab.”

Fehrenbacher’s group noted that atezolizumab “significantly improved survival compared with docetaxel in patients with previously treated NSCLC. Improvement correlated with PD-L1 immunohistochemistry expression on tumor ce/p>lls and tumor-infiltrating immune cells,” leading them to conclude: “PD-L1 expression is predictive for atezolizumab benefit. Atezolizumab was well tolerated, with a safety profile distinct from chemotherapy.”

The researchers performed the clinical trial using immunohistochemistry, and also ran a gene expression panel of 91 genes featured in the Herbst paper. These genes are included in the Advanta Immuno-Oncology Gene Expression Assay. The team subsequently used these results to develop a gene signature based on a subset of the panel, effectively predicting therapeutic response that provides data that could potentially be used for developing future diagnostics tests.

Achieve greater overall productivity

The Advanta Immuno-Oncology Gene Expression Assay is optimized to run on the industry-proven Biomark HD system. This automated system enables researchers to easily run up to 96 samples at once with limited hands-on time, while delivering trusted performance. Together with the capability to sensitively detect biomarkers across defined T cell subsets, immune regulation, immune cell fate, cytokines, chemokines and more with exceptional cost-efficiency, this new assay offers an ideal solution to meet the rigorous demands of translational research studies.

Advanta IO Gene Expression Assay ordering information

Contact us to learn more about the Biomark HD system and the Advanta Immuno-Oncology Gene Expression Assay.

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For Research Use Only. Not for use in diagnostic procedures.

Enabling multiparameter profiling of precious samples, mass cytometry has been instrumental in achieving new research breakthroughs in areas such as immunology, cancer immunotherapy, drug discovery, vaccine development and more around the world.

Watch the webinars below to learn how researchers are utilizing Helios™, a CyTOF system, and mass cytometry applications to further their research.

	Finding Pathologic T Cells in Rheumatoid Arthritis by Mass Cytometry Deepak Rao, MD, PhD  Rheumatologist Co-Director, Human Immunology Center Brigham and Women's Hospital Watch
	Querying Cytokine-Signaling Networks in Myeloproliferative Neoplasms Stephen Oh, MD, PhD  Assistant Professor of Medicine, Division of Hematology  Washington University School of Medicine  Watch
	Enhancing Checkpoint Blockade in Lymphoma with In Situ Vaccination Joshua Brody, MD  Assistant Professor, Hematology and Medical Oncology Director, Lymphoma Immunotherapy Program Icahn School of Medicine at Mount Sinai Watch
	Decoding Tumors with Mass Cytometry Jonathan Irish, PhD Assistant Professor of Cancer Biology, Assistant Professor of Pathology, Microbiology and Immunology Vanderbilt University Watch
	Defining the Phenotypic and Functional Immune Microenvironment in AML Evan Lind, PhD Assistant Professor, Department of Molecular Microbiology and Immunology and Department of Cell, Development and Cancer Biology Oregon Health & Science University Watch
	Towards an Assay of Global Immune Competence Holden Maecker, PhD Associate Professor (Research) of Microbiology and Director, Human Immune Monitoring Center Stanford University Watch
	Signatures of Protective NK Cell Responses Catherine Blish, MD, PhD Assistant Professor, Department of Medicine and Immunology, Stanford University School of Medicine Assistant Director of the Stanford Medical Scientist Training Program Watch

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For Research Use Only. Not for use in diagnostic procedures.