Spatial phenotyping is the power to visualize and quantify dozens of biomarkers in a single tissue sample while maintaining cellular and sub-cellular detail. Comprehensive spatial phenotyping can reveal cell diversity, co-expression patterns, cellular interactions, broader tissue architecture, and much more – all from a single tissue section.
It takes an integrated systems biology approach to uncover insights and make the unexpected connections that ultimately lead to true breakthroughs. The systems used to conduct spatial phenotyping experiments must be capable of spatially resolved, highly multiplexed biomarker analysis. Our proven solution for conducting unbiased spatial phenotyping studies has long been the PhenoCycler™ (formerly CODEX) solution and now ushering in a new era is the PhenoCycler-Fusion.
The PhenoCycler-Fusion is an integrated platform combing the strengths of Akoya’s automated, ultrahigh multiplex cycling platform, PhenoCycler, and its high-speed imaging platform, PhenoImager™ (formerly Phenoptics), into an end-to-end, integrated workflow. These newly evolved capabilities will transform how we study tissue biology. Here we present 10 reasons why we are excited by the prospect of PhenoCycler-Fusion and how we expect it to be a game changing technology in the field of spatial biology.
In the old world, researchers conducting a spatial biology experiment had to deal with an iron triangle – a concept by which resolution, plex, and throughput could not all be simultaneously improved.
For example, single-cell technologies, such as flow cytometry and single-cell RNA-seq, can detect numerous parameters but lack spatial dimension. The advent of recent spatial technologies has alleviated this concern, but spot-based or region-of-interest-based methods still lack resolution. Standard imaging methods, like immunofluorescence (IF) and immunohistochemistry (IHC) provide single-cell resolution and spatial context but are limited to measuring just a few parameters simultaneously.
Akoya has developed a way to break this iron triangle – by the virtue of speed. The PhenoCycler™-Fusion system is the fastest single-cell spatial biology platform on the market. The workflow is about 10 times faster than workflows involving standard microscopes. With PhenoCycler-Fusion we can achieve sample-to-data within 24 hours.
There are 4 basic components to a spatial phenotyping experiment: 1) tissue prep and staining, 2) imaging, 3) data transfer and processing, and 4) data analysis. To cut down on tissue prep and staining, Akoya has developed pre-designed antibody panels (custom panels are also an option) which are validated and experiment ready. To reduce imaging times, Fusion enacts high-speed, state-of-the-art imaging coupled with automated cycling to scan 1 million cells in 10 minutes, benchmarking it as the fastest spatial biology system available.
2. Tunable workflow
PhenoCycler-Fusion puts you in the driver’s seat. PhenoCycler-Fusion will serve the entirety of the spatial biology continuum, allowing fine tuning of your workflow to match your customized needs. The scalable nature of the PhenoCycler-Fusion system allows large high-plex panels for unbiased discovery or focused panels emphasizing high-throughput and sample size when in the translational or clinical stages of your research.
3. Walk-away automation
The Fusion has been outfitted with several sophisticated capabilities allowing it to be user-friendly and automated. Among those capabilities are its high-speed optics and stage that move quickly and smoothly in the x, y, and z directions, this is what focuses the tissue for imaging rather than the tediously adjusting of optics. This process is completely hands-free, providing ultraprecise focusing and maximizing the efficiency of multiple downstream steps.
The Fusion is also capable of automatic tissue detection (focal point detection and alignment) prior to imaging. This takes the guesswork out of configuring your microscope before image capture. With PhenoCycler-Fusion you simply place the slide in the flow-cell carrier and let the instrument take care of the rest.
We understand how precious tissue biopsies can be and the importance of protecting this resource when performing an experiment (see reason 5). It’s important to know in advance that a spatial biology solution will be compatible across tissue types. PhenoCycler-Fusion can successfully stain and image virtually any sample type including but not limited to FFPE tissue, fresh-frozen samples, archived whole slides, tissue microarrays, breast, brain, lung, spleen, kidney, lymph nodes, skin, and organoids.
To maintain its high-speed capabilities, the PhenoCycler-Fusion cannot have any wasted movements, reagents, data, or tissue. The sophisticated automated focusing capabilities (discussed in reason 3) use ultraprecise movements during the imaging phases. These fine-tuned movements allow seamless focusing of slides and optimization of light exposure resulting in rapid image acquisition with high resolution.
The PhenoCycler-Fusion is uniquely capable of imaging non-rectangular shapes – leading to efficient imaging and coverage of the tissue. The novel flow-cell carrier integrates the fluidics of PhenoCycler with the stage of Fusion while moderating the flow of reagents across the tissue. In doing so it ensures that reagents are used with optimum efficiency, and the exact prescribed amount of reagent is used throughout each run, reducing waste and ensuring effective hybridization and dehybridization of barcodes and reporters. PhenoCycler-Fusion’s attention to detail at every step means more tissues being imaged, more data being generated and analyzed, and more actionable insights.
6. A foundation of proven technology
Many high-impact peer-reviewed journals, including Nature Communications, Cell, and Science, have demonstrated that the PhenoCycler and PhenoImager technologies are essential for comprehensive spatial phenotyping. The number of publications involving Akoya’s spatial biology instruments has increased 10-fold in just 2 years, going from 27 publications in 2019 to over 260 publications in 2021.
Of note, several novel and interesting applications, including coordinated cellular neighborhoods that reveal spatial organization of the tumor microenvironment , patient stratification using SpatialScores  identification of epithelial subpopulations that are predictive of therapeutic response  and deep spatial phenotyping revealing hierarchical structural networks , have paved the way for spatial discovery and actionable insights. With the power and speed of PhenoCycler-Fusion, we expect the pace of discovery to accelerate even faster.
A whole slide human FFPE sample with more than 3 million cells, imaged on the PhenoCycler-Fusion.
8. Fusion is multiomic
Akoya has made a commitment to truly define a wide range of spatial applications – one application is defining spatial functional states. Spatial functional states are how cellular phenotypes interact with each other – which pathways are turned on and which are turned off. To garner a deep and complete understanding of spatial functional states we must approach these types of studies from a multiomic perspective. Fusion is well-suited for the task.
The PhenoCycler-Fusion system combined with Bio-Techne’s proven and established RNAScope® technology will enable comprehensive spatial phenotyping of RNA and protein biomarkers. The synergy of RNA and proteomics is extended across one platform and we continue to explore and demonstrate different RNA chemistries while utilizing well-established protein biomarker assays for complete RNA and protein expression mapping.
9. Supported by a passionate team
PhenoCycler-Fusion is supported by a passionate team of Akoyan’s with extensive expertise such as technical specialist and instrument engineers to assist in day-to-day operations as well as experimental analysis. In addition to in-person resources we also offer freely available videos and documents that guide users through the PhenoCycler-Fusion workflow. The PhenoCycler-Fusion Support team serves as an expert resource to actively guide users through installation, training, and beyond.
PhenoCycler-Fusion users are encouraged to attend virtual Akoya Academy training sessions to share tips, tricks, and best practices on using PhenoCycler-Fusion technology and create an inclusive and collaborative environment. Akoya Academy training topics include custom panel development, conjugation best practices, image optimization, data analysis pipelines, and more. PhenoCycler-Fusion users also get exclusive access to user-group meetings and workshops to encourage sharing best practices and success stories across the community. We ultimately hope these resources help users to minimize disruptions in productivity and maximize performance in the lab.
10. Fusion is fueling the next wave of discovery biology
Spatial researchers agree that multiomic technologies are going to be far more widely used in the future. We are seeing the need for and the advent of analytical frameworks and algorithms to integrate imaging-based single-cell spatial phenotyping data with complementary transcriptomic and genomic datasets. High-plex cell phenotyping methods like single-cell RNA-seq capture the deep cellular heterogeneity of samples, but cell behavior is a function of all that surrounds it.
Imaging-based spatial phenotyping platforms enable researchers to visualize and analyze cell diversity, interactive networks, and cellular behavior across whole tissue sections. Both types of data have complementary features and now, for the first time, PhenoCycler-Fusion fuses information about a cell’s proteome and transcriptome with single-cell, spatial context.
Ready to take your spatial biology workflow to the next level? Discover PhenoCycler-Fusion.
- Schürch et al. Coordinated Cellular Neighborhoods Orchestrate Antitumoral Immunity at the Colorectal Cancer Invasive Front. 2020, Cell 182, 1341–1359.
- Philips, et al. Immune Cell Topography Predicts Response to PD-1 Blockade in Cutaneous T Cell Lymphoma. Nat Commun. 2021 Nov 18;12(1):6726.
- Gouin, K.H., et al. An N-Cadherin 2 expressing epithelial cell subpopulation predicts response to surgery, chemotherapy, and immunotherapy in bladder cancer. Nat Commun 12, 4906 (2021).
- Hickey, J.W., et al. High-Resolution Single-Cell Maps Reveals Distinct Cell Organization and Function Across Different Regions of the Human Intestine. bioRxiv Nov 2021 469203.