2018 ISS Abstracts • •
The Industrial Science Symposium will take place Friday evening, September 28th. This session is intended to provide our exhibitors, a forum to present new technology, instrumentation, etc. Each presentation lasts approximately 15 minutes.
Nanostring Technologies Inc.
Validation of antibody panels for high-plex immunohistochemistry applications
Indira Medina (firstname.lastname@example.org)
Douglas Hinerfeld, Kristi Barker, Heather Metz, Chris Merritt, Lucas Dennis, Philippa Webster, Indira Medina, Joseph Beechem
Introduction: Characterization of the spatial distribution and abundance of key proteins and cells within tissues enables a deep understanding of biological systems. However, it has proven difficult to perform such studies in a highly-multiplexed manner on FFPE tissue sections. There has been significant progress in developing technologies with expanded capabilities to analyze higher numbers of proteins, however, the validation of these technologies and their associated affinity reagents remains a significant barrier to adoption. We have developed a validation pipeline that ensures optimal sensitivity and specificity for high-plex antibody panels for the analysis of FFPE sections using the NanoString Digital Spatial Profiling (DSP) platform. The DSP is designed to allow visualization and detection of cells in FFPE sections, and simultaneously analyze up to 96 proteins using antibodies conjugated to oligonucleotides that can be released via a UV-cleavable linker for quantification. Methods: Antibodies targeting immuno-oncology proteins were tested for specificity and sensitivity by immunohistochemistry on FFPE human tissues, as well as human cell line pellets to evaluate binding specificity of both unconjugated and oligo-conjugated antibodies. The sensitivity and dynamic range were tested using FFPE cell pellets with target-specific positive and negative cells at different ratios. An interaction screen was performed to evaluate potential deleterious effects of multiplexing antibodies, and a human tissue microarray (TMA) containing normal and cancer tissues was employed to assess assay robustness. The reproducibility of the panel on DSP was tested on serial FFPE tumor specimens by correlating the expression of all markers across 24 spatially-registered regions of interest (ROI) as well as the ability to reveal biological heterogeneity within lymphoid tissue by characterizing the expression of 40+ proteins in a spatial grid of 100um x 100um ROIs. Results: Immunohistochemical analysis of unconjugated and oligo-conjugated antibodies displayed indistinguishable staining patterns on control tissues and cell lines. Mixed cell pellet assays revealed strong correlations between observed counts and positive cell numbers. Antibody interaction studies showed similar count values for antibodies alone or in combination, and TMA hierarchical clustering analysis demonstrated expected patterns of expression across tissue types. Analysis of all markers across 24 registered regions of interest across serial FFPE sections were highly correlated. Spatial analysis of lymphoid tissue revealed high levels of biological heterogeneity across multiple germinal centers. Conclusion: These results demonstrate the validation and application of high-plex protein panels to accurately interrogate the immune biology within FFPE tissue using the NanoString DSP platform.
Streamlined human immune monitoring with mass cytometry: 29 markers in a single tube and automated data analysis
Michelle M Poulin (email@example.com)
Immune monitoring is an essential method for quantifying changes in immune cell populations in health and disease. The extreme heterogeneity of immune cells demands a high-parameter approach to more fully and efficiently quantify these changes. Mass cytometry is an ideal solution, enabling simultaneous detection of over 40 phenotypic and functional markers in a single tube of sample.
We developed a single-tube, 29-marker panel for mass cytometry based on the Human ImmunoPhenotyping Consortium (HIPC) consensus panel (Maecker et al. Nature Reviews Immunology, 2012), expanded to allow identification of additional leukocyte subsets, particularly T cells. Automated data analysis with Verity Software House GemStone™ has been developed specifically for data collected with the panel providing users with results in minutes, reducing time-to-answer and variability inherent in manual gating.
We will describe the kit and available data analysis options, as well as presenting data from extensive internal testing of the kit for repeatability, reproducibility and panel performance comparing population frequency data using the full 29-marker panel to that obtained with several 10-12 marker panels.
Union Biometrica, Inc.
COPAS VISION flow cytometer captures images and sorts
Yongwoon Kim (firstname.lastname@example.org)
We have developed instrumentation for large particle flow cytometry that can capture brightfield images in flow. Adding imaging capability to flow cytometry greatly enhances the phenotyping of samples by providing morphological and spatial information of the sample constituents not collected by conventional flow cytometers. Traditional measurements of size, optical density, and fluorescence, as well as Profiler data, are also collected, and these measurements are used for making sorting/dispensing decisions. The collected images and flow cytometry measurements are synchronized so that objects dispensed to wells of multiwell plates can be traced back to their corresponding image. Our COPASTMtechnology platform is designed for large particles making it ideally suitable for large single cells, cell clusters, and small model organisms. The COPASTMVISIONinstrument is based on this platform and is ideally suited for samples made up of particles of varying sizes and shapes. Our data from the COPAS VISIONshows proof-of-principle support for increased level of phenotyping of these types of samples.
Using FCS Express to Accelerate your Flow Cytometry Reporting and Results
Sarah Schuett (email@example.com)
Most flow cytometry data analysis software focuses on generating plots, gates and stats. But those are rarely your final results. Most researchers spend a significant amount of time transferring data from their flow analysis software into other packages (like Excel, Prism, Jump, Powerpoint and many others) for downstream processing. This dramatically increases the time it takes for you to get your results. FCS Express 6 has a wide variety of tools to bring your final results to you directly in your flow analysis software. Integrated bar and pie charts update instantly as you change your gates. A fully functional spreadsheet lets you apply sophisticated calculations and regressions that change automatically with your analysis. Direct R integration as well as SPADE and vSNE plots provide state of the art high dimensional analysis and visualization. New index sorting compatibility allows for quick single cell and well review at a click. Come see how FCS Express can help you produce your results in record time.
Todd Salomon (todd.Salomon@emdmillipore.com)
Discover unparalleled fluorescence sensitivity and flexibility in a compact and affordable flow cytometry system. With patented Amnis® optics technology inside, the new CellStream™ flow cytometer uses a camera for detection to rapidly capture low-resolution cell images and transform these into high-throughput intensity data. Whether you are looking for small particles (200nm) or feint signals, the CellStream™ flow cytometers can be fully configured with 1 to 7 lasers and are easy to upgrade directly in the laboratory and with your applications in mind.
Expanding Application Capabilities Using Full Spectrum Cytometry
Dave Kennedy (firstname.lastname@example.org)
The Aurora continues its development path with expanded capabilities. Data will presented with the addition of 561 laser. Panel data with 4 lasers, 20+ colors. Examples of autofluorescence extraction, now enhanced with 2.0 software.
Novel Aspects of Sorting in a Closed System:The MACSQuant® Tyto®
Dr. Patrick Sean Murphy (email@example.com)
The MACSQuant® Tyto® is a revolutionary microfluidics based cell sorter allowing for gentle, high purity cell isolation in a closed, sterile environment. Designed around the world’s fastest mechanical valve, the Tyto® enables sorting of fragile and hard-to-isolate cell types without high-pressure or charge, resulting in unparalleled post-sort viability and functionality. The unique sort mechanism and disposable cartridge completely eliminates aerosols and the risk of sample cross-contamination, making Tyto® an ideal choice in highly regulated environments. Applications regularly demonstrate >90% purity while maintaining >95% viability, even with delicate and rare populations such as iPS cells and mesenchymal stromal cells. The MACSQuant® Tyto® represents the state-of-the-art in closed system sorting, providing full sterility, safety and simplicity for cell isolation.
Sony Biotechnology Inc.
Multi-application sorting with the Sony MA900
Mr. Steve Whitaker (Stephen.Whitaker@sony.com)
The MA900 is a precision benchtop cell sorter that is easy to use, supports 12-fluorescence parameters, 4-way sorting and single cell deposition into multiple plate types. Use of modern technologies and integrated automation dramatically simplify operation to make sorting less subjective and improve reliability.
System startup, aseptic cleaning, QC, and sort setup operate with a touch of a button to ensure optimal daily alignment of the sorting chip to lasers, precise targeting, and rapid recovery from clogging. Guided workflows for cleaning cycles simplify maintenance and can be customized for aseptic sorting. Versatile system design provide capability to sort a wide range of applications ranging from single cell sorting to multicolor immune cell panels.
Sorting Simplified: Introducing the BD FACSMelody
David Morris (firstname.lastname@example.org)
In this presentation, we will introduce the audience to the BD FACSMelody system. We will show how the simplified operation of the system brings truly easy cell sorting to shared research core facilities and individual laboratories. The FACSMelody combines proven and exclusive BD technology found in our high-end sorters with new automation, simplified software interfaces, and guided workflows. This means researchers are empowered to set up and optimize their own sorts in a fraction of the time it takes to set up other sorters. The standard 100 mm nozzle in the FACSMelody gives researchers flexibility to efficiently sort bacteria, yeast, lymphocytes, or mammalian cell lines. Equipped with optical filters optimized for the brightest commercially available dyes, the FACSMelody facilitates the separation of rare populations using up to nine fluorochromes.
Page updated on 2018-08-08 19:59:20 -0400