Imaging Mass Cytometry Services

Visikol has established a partnership with Fluidigm through which it is now offering imaging mass cytometry (IMC) services to its Clients such that any researcher can now image their tissues using more than 30 markers for a single slide. The addition of this IMC capability extends Visikol’s existing fluorescence multiplexing capability and now provides researchers with the ability to survey a larger number of markers from a single slide while greatly minimizing background noise and the challenges associated with markers that have low expression.

What exactly is IMC?

IMC is a relatively new technology that has been pioneered by Fluidigm with their Hyperion™ Imaging System and many researchers in the tissue analysis space might not be yet aware of this technology platform. Typically, when we try to image a slide, we are using either chromogenic staining or fluorescent staining in conjunction with brightfield or fluorescent slide scanning wherein we are detecting the presence of a marker through the use of selective stains and imaging. These approaches are highly reliable and high throughput but are typically limited in the number of markers that can be acquired at one time from a single slide. In the case of fluorescent slide scanning, we can acquire 4-5 channels from a single slide at a time, but to move beyond this we need to optimize a stripping and relabeling process which can be challenging to implement and validate. Furthermore, fluorescence imaging is prone to fluorescence overlap and challenges with background fluorescence that can minimize image quality.

IMC was developed to allow for a high level of multiplexing wherein more than 30 markers can be obtained simultaneously from  a single slide. The principle of IMC is very different from traditional slide labeling and imaging whereas there is actually no camera used to image the slides. Instead, slides are labeled with antibodies that are conjugated to metals and a 1 µm ablation laser is used to ablate a 1 m square of tissue off of a slide at a time whereas the resulting metal ions are fed through a mass spec to analyze the metal ions present and their respective quantity. Through moving the ablation pixel across a region of interest on the slide, a 1 mm area of tissue can be analyzed in approximately two hours for up to 40 markers. The mass spec data from each pixel can then be used to create an image of the region of interest that was ablated such that mass spec data can be converted into an image for traditional image analysis.

What are the advantages of IMC?

Because IMC uses metal tagged antibodies instead of fluorescence, there is no overlap between peaks and no background noise as the metals used do not naturally occur in nature. Therefore, IMC image data sets are very crisp and clear and have a very high degree of sensitivity when compared to fluorescent imaging. The ability to multiplex up to 40 markers at the same time means that tissue processing is simple and straightforward, and a large number of targets can be profiled simultaneously.

What are the disadvantages of IMC?

Like every imaging tool, there is a specific use case in which IMC is best adapted and we find that to be for research questions where large numbers of targets need to be studied simultaneously (e.g. immune-oncology) and where targets might have a very low expression profile that could bleed into the background with traditional fluorescent imaging. The reason that IMC cannot be used for all research questions is that it is limited in the amount of tissue that can be surveyed as it takes two hours to image a 1 mm square from a tissue and the generated data set is only approximately 10X magnification. This is compared to fluorescent slide scanning where we can image five channels from a whole slide at 40X magnification in 15 minutes and achieve about 4,000 times more data in the same amount of time. Additionally, IMC is expensive as it requires a lot of consumables (e.g. argon, antibodies) to image a single region of interest.

What IMC services does Visikol provide?

Visikol provides Clients with end-to-end tissue-to-insight services where Clients can ship Visikol tissue samples and receive back a detailed report that provides a quantitative assessment of each individual slide and a report addressing the Client’s specific research question. These services include the following:

• Traditional tissue processing (i.e. embedding, sectioning).
• Custom label panel development and optimization.
• Ablation and image generation.
• Image analysis and reporting.

How we conduct IMC projects

1. Visikol and the Client have a detailed conversation to discuss the project scope and the desired end-points.

Typically, most of our Clients are interested in more than just raw image files and we work closely with our Clients to understand their specific research question and how we can quantitatively address their question using our suite of image analysis software. From this conversation, our team prepares a quote and statement of work for these efforts.

2. The Client then sends to Visikol either tissue blocks for slide prep or slides for labeling.

3. Visikol leverages an existing panel of labels or optimizes and validates a panel specifically for the Client and then labels the slides.

4. Prior to imaging, Visikol works with the Client to select regions of interest from each slide for ablation.

5. The slides are ablated and the raw image files are generated.

6. Visikol uses these raw image files to generate a customized report for the Client. These range in complexity from simple cell counts to much more complicated analyses dependent upon the Client’s research question.

What validated panels does Visikol have?

Visikol has access to several off-the-shelf antibodies and panels which can be used in Client projects. Additionally, Visikol can develop and validate custom antibody panels and conjugate new antibodies that it does not yet offer. Visikol currently offers all of the standard Fluidigm Maxpar panels as well:

• Maxpar Human Tumor-Infiltrating Lymphocytes IMC Panel Kit
• Maxpar Human Immune Activation IMC Panel Kit
• Maxpar Human Tissue Architecture IMC Panel Kit
• Maxpar Catalog