• Scanned images generated on the Leica Aperio AT2 can be opened using Leica’s ImageScope software (Windows) or QuPath (iOS).
• Scanned images generated on the Zeiss Axio Scan.Z1 can be opened using Zeiss ZEN Lite (Windows) or or QuPath (iOS).

If you are new to ZEN, please check our Quick Start Guide!

Yes- project consultation can be requested through the service request form. Please select ‘Consultation’ in question 1 and continue filling the remainder of the survey.

Majority of ROI’s/AOI’s are selected one of 2 ways:

1. Morphology mask guided. Each slide is stained and scanned for 4 immunofluorescence biomarkers. These markers are used to autogenerate segmented regions based on their expression. For example, Pan-cytokeratin would be used to generate the tumor segmented region.
   • Ready to use morphology kits are available to be purchased by the researcher from Nanostring.
   • Custom masks/antibodies can be optimized by MAPcore on an individual bases for a fee.
2. User guided. Researchers can request to be on-site during the DSP protocol to select the ROIs themselves.

For more comprehensive explanation of ROI selection please click the link to the DSP manual.

The number of ROIs per project is dependent on the assay being run.

Please see the slide planning guide for further information.

Multiplex panels come in 2 flavors- brightfield/chromogenic and fluorescence.

• Brightfield multiplex panels currently have the capacity of 4 biomarkers plus hematoxylin (5 colour). However, depending on panel composition, more biomarkers can be added. Please note that multiplex chromogenic panels must be scored by-eye by a pathologist and cannot be scored using automated software.
• Fluorescence multiplex panels have capacity for 7 biomarkers plus DAPI (8 colour). These panels can be scored using automated software by MAPcore (additional charges apply) or by researchers using digital pathology software of their choice (ex. QuPath or Halo).

For each provided service, MAPcore will typically provide majority of reagents. However, researchers will need to to provide the below material for each service requested:

No. The GeoMx nCounter assays have been developed with unique UV-cleavable probe sets. However, 10 custom probes can be added to any nCounter GeoMx DSP panel. If you are interested in adding custom probes please contact us for more information.

Please explore the currently available probes lists from NanoString for the GeoMx DSP:

Multiplexed cyclic immunofluorescence allows analysis of multiple biomarkers and their spatial relationships within a tissue. In each cycle, the tissue is stained and imaged with a set of up to four antibodies. After imaging, the dyes are inactivated, enabling repeated rounds of staining and imaging on the same sample.

Image credit: https://www.leica-microsystems.com/science-lab/life-science/spatial-biology/be-confident-in-your-results-with-cell-dive-validated-antibodies/

FFPE tissue samples are first dewaxed, then undergo sequential antigen retrieval using an acidic citrate buffer followed by an alkaline EDTA buffer. This process reverses crosslinking and exposes a broad range of epitopes for staining and imaging.

Each sample will then go through rounds of:

1. Autofluorescence imaging  

2. Biomarker labelling 

3. 20x Biomarker imaging (488/555/647/750 LEDs) 

4. Dye inactivation 

Dye inactivation is achieved by breaking the methine bridge of cyanine-based dyes through oxidation with hydrogen peroxide. Autofluorescence imaging after each inactivation round serves as a quality control check, ensuring no residual signal from previous staining rounds persists. To avoid species cross-reactivity, we recommend using antibodies that are directly conjugated to one of these fluorophores: Alexa Fluor 488, Alexa Fluor 555, Alexa Fluor 647, Alexa Fluor 750

We have had the most success with FFPE tissues and TMAs sectioned at 4um thickness. Extensive testing has not been done with fresh frozen samples; however, this should theoretically be possible with thin sections less than or equal to 10um thick. 

The maximum mounting area for tissues is 4cm length x 2cm width

Cell DIVE collects images with a 20X objective at 2040 x 2040 pixels for each field of view. The pixel resolution is 6.5um squared per pixel. 

Leica claims the ability to detect over 60 markers on a single tissue sample. Although we have not yet fully validated this claim in our hands, we have successfully performed up to six rounds of dye inactivation and staining (up to 6×4=24 antibodies!). Remarkably, we have not observed any significant antigen degradation or epitope loss in the tissues. 

Before each imaging session, autofluorescence images are compared to the baseline to detect any shifts in position or rotation. These adjustments are then applied to the slide’s coordinate system to ensure consistent alignment across layers. The DAPI image from the autofluorescence round serves as the reference for algorithmic registration of subsequent layers. This approach aligns images based on the entire cellular content of each field of view, rather than relying only on overlapping edges used in stitching. 

Please contact us (map.core@ubc.ca) for the most recent copy of validated antibodies  

We recommend placing markers of outmost importance or interest in the first few rounds of the experiment, with explorative markers placed in later rounds of the experiment. Markers that are weakly expressed in the tissue and or secreted proteins that are known for high background should be paired with Alexa Fluor 647 or 555. Markers that are highly expressed/strongly stained should be paired with Alexa Fluor 488 or 750. 

As a general guideline, the signal-to-noise ratio typically follows this pattern: 

Alexa Fluor 647 > 555 > 750 > 488. 

It is also possible to optimize antibody concentrations through repeated staining of the same antibody across rounds. 

Yes, MapCore works closely with the UBC Ablab to conjugate antibodies, we have experienced proven success with antibodies that are custom conjugated on the CellDive platform. 

Antibodies will need to be carrier free (BSA free, preferably azide free as well) in order to not interfere with conjugation chemistries. 

Yes, MapCore has access to AIVIA AI image analysis software, equipped with machine learning-powered nuclei segmentation tools, as well as advanced features like unsupervised clustering and algorithms for determining closest object relationships.

Example of segmented cells with AIVIA software: