Imaging Mass Cytometry Service

Imaging Mass Cytometry Service2020-10-15T19:45:01-05:00

Background

  • Imaging Mass Cytometry (IMC), also known as CyTOF imaging, utilizes principles of mass cytometry to generate images with a wide range of antibody panels and offers simultaneous characterization of many antibody markers in a single scan. As in mass cytometry, antibodies are conjugated with isotopically pure metals to label cellular proteins. Tissues are then ablated using 1 um square pixels and the metal-conjugated antibodies are ionized following passage through an inductively coupled plasma to be analyzed by mass spectrometry [1]. Usage of metal-conjugated antibodies overcomes limitations of traditional fluorophore-based cytometry techniques, which can have a broad range of emission spectra contributing to noise. The technique also allows for multiplex immunophenotyping with high-dimensional cellular analysis of up to 40 labels at once [2], granting a much deeper understanding of biological models and the interrelationships that contribute to tissue processes.
  • Using mass spectroscopy, images can be created with the data from each subsection of tissue for all labeled markers. By analyzing small discrete areas at a time (usually 1 um square subsections), noise is reduced as each isotopically pure metal should theoretically have distinct detection peaks. As such, imaging mass cytometry (CyTOF) is an excellent way to produce high content images of tissue structures while minimizing many of the noise related issues that come with using other imaging modalities.
  • With our suite of quantification and image analysis services, and recent partnership with Fluidigm, Visikol has become a leader in the IMC imaging and image analysis space. As experts in high content 3D imaging, our team of scientists can help provide quantitative insights for your next experiment.

Protocol

InstrumentHyperion Imaging System
Analysis MethodImage Processing
MarkersVariable, up to 40 total markers during one run, catalog available upon request
Sample SubmissionWhole Tissue or Tissue Blocks, Digitized Image Slides, Processed Images
Imaging Parameters10X magnification with 1 mm2 regions of interest
Quality ControlsHistogram intensity analysis of negative, positive controls
Data DeliveryROI masks of detected cells overlaid on slide images, data tables containing cell counts for all cell types, histograms of spatial distribution for nearest neighbor analysis, statistical analysis comparing results between regions or different slides, other quantification strategies available on request

General Procedure

  1. Tissue sample is transferred to Visikol
  2. Custom label and marker panel is developed for Client specifications
  3. Sample is processed, sectioned, labeled, and imaged using the Hyperion Imaging System
  4. Resultant images are processed and analyzed according to Client specifications
  5. Images and quantification report is generated and transferred to Client

Data

Representative Segmentation Data

Figure 1. Sample segmentation protocol for visualization of Ki67 tumor cells within a slice of labeled tissue (using Erbium 168)

Representative Nearest Neighbor Analysis

Figure 2. Visual representation of nearest neighbor analysis for assessment of CD3 positive cell distance (T-cells) from tumor edge.

Representative Overlay Examples of Multiple Markers

Figure 3. Sample overlay of multiple markers in a single image for visualization of signal density

Representative Quantification of Image Data

Figure 4. Bar graph depicting distribution of cell types in tumor vs non tumor tissue

Figure 5. Bar graph depicting spatial distribution of CD3 T-cells within tumor tissue (check figure 2 for image representation)

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