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Analysis of immunohistochemical markers

HistoQuest provides four analysis modes:

  • Cell-based (nucleus, cytoplasm, membrane)
  • Stained areas
  • Small dots (e.g. CISH)
  • Skeleton algorithm for membrane staining

Context-based analysis can be done by manually drawing regions-of-interest. TMAs can be analysed and detected on imported files.

HistoQuest has importers for the following digital image or slide formats:

  • 3D Histech (.mrxs)
  • Hamamatsu (.ndpi)
  • Zeiss Axio Scan (.czi)
  • Leica (.scn)
  • Aperio (.svs)
  • Images (.jpeg, .tif, .bmp, .png) 
  • Generic Big Tiff
  • OME-Tifff

is integrated into the following TG systems:

  • TissueFAXS SL
  • TissueFAXS SL PLUS
  • TissueFAXS Q+
  • TissueFAXS PLUS
  • TissueFAXS Histo
  • TissueFAXS i PLUS
  • TissueFAXS i Histo

HistoQuest is also available as a standalone software.

HistoQuest 6.0

HistoQuest is a brightfield image analysis software for the FACS-like analysis of samples stained with immunohistochemical or histochemical stains.


NEW FEATURES in HistoQuest 6.0

HistoQuest 6.0 comes with new features which improve handling of large samples/number of samples in one project as well as increase the softwares speed:

  • Full 64 bit support will greatly increase the number of samples which can be analysed in one project.

  • The latest processing libraries have been integrated which will increase processing time by up to 40%.

  • The cache from TissueFAXS projects will be used, this eliminates wait time while the project is loaded into the viewer.

  • Finally, HistoQuest has a heatmap feature in scattergrams. This feature will show areas of objects density in the scattergram which may not be visible in highly populated scattergrams and so deliver more information when working with them.


HiQ6 heatmap


TissueGnostics introduced Tissue Cytometry technology into image analysis software more than a decade ago. While, at a first look, it may appear to be just another form of displaying data, there is much more to it. Combined with segmentation images it is also a simple and intuitive tool to visually validate results. TG calls this "Backward Gating". Backward gating can be done for any measured parameter on histograms and scattergrams. The following examples demonstrate the versatility of backward gating. In the left image backward gating is used to visualise cells in which the HER2/neu staining has a closure angle of more than 180 degrees. In the right image backward gating is used on segmented nuclei positive for Ki-67. 

Backward gating Histoquest

Morphological filtering using Tissue Cytometry technology combined with appropriate planning of the analysis provides a highly flexible and capable instrument for extracting specific data from a sample (see image below).

Tissue Cytometry technology

HistoQuest analysis automation

HistoQuests fully automated template-based analysis is optimised for clinical routine and research applications and allows complete walk away operation.

The user just has to load samples into the software and to decide whether they want the samples to beautomatically detected (in which case analysis will start immediately), whether they want to be able to interfere with the automatic region detection (with validation) or whether they want to manually define regions-of.interest.


A template provides:

  • Segmentation settings for all objects to be detected
  • A method for dealing with events in ROI
  • Markers
  • Diagrams from the first sample of the project from which the template was saved
  • Statistics report settings
  • Simple report settings
  • Small dots settings 



HistoQuest image analysis algorithms

HistoQuest AlgorithmBrightfield images need to be separated into images according to their color components, HistoQuest provides two algorithms for color separation. 

The first one automatically detects and separates two colors and an optional third one. The second color separation algorithm is semi-manual (using a color picker) and separates an unlimited amount of colors with great control and precision.

HistoQuest uses algorithms for nuclear and cell compartment segmentation. It also provides an algorithm for stained area detection as well as algorithms for the detection of dots inside of already detected cell compartments.




The nuclear segmentation algorithm is highly efficient in difficult situations in tissue and can be set with two values only - mean nuclear size and nuclear channel background threshhold. The cell compartment algorithm provides a large amount of flexibility with a few simple settings (yellow outline = segmented nuclei, blue outline = nuclei positive for DAB or cytoplasm positive for Permanent Red marker). The total area algorithm segments stained areas in a sample.


 The dot algorithm provides small dot detection ideal for CISH, ISH and other applications in which the quantification of small objects within cellular compartments is needed. The images below show the quantification of EGFR RNA dots inside of the nuclear compartment. Simple context-based analysis can easily be done in HistoQuest by using manual region tools. Drawing the regions takes a negligible amount of time and provides data for epithelial areas and stroma separately in this example. The skeleton algorithm permits the detection of membrane stainings.

quantification of FITC and Texas Red FISH dots inside of the nuclear compartment.


Analysis Support

HistoQuest fully supports the analysis of Tissue Microarrays (TMA) scanned either with TissueFAXS systems or those slide scanners for which HistoQuest provides importers. For analysis projects of scans coming from TissueFAXS systems the autodetected TMA regions will be imported. For scans imported from other scanning systems without detected regions the TMA autodetection feature is also available in HistoQuest.


HistoQuest Data Export capabilities

TQ export2HistoQuest provides four modes for the export of analysis data, ranging from simple prefabricated reports for internal presentations to selective data export from all analysis projects from a complete scientific project.

There is a special pathology-oriented data export feature for the small dot algorithm data, which has been built according to pathologists requirements for FISH analysis.

All TG data export modes write data to Excel, CSV and PDF formats.



Other HistoQuest features

HistoQuest features include a registration and comparative analysis capability for differently stained consecutive samples. Compare sets provide grouping features for multiple scattergrams and histograms as well as overlaying them.

TQ export3


tg features


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