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HALO® Module

ISH-IHC Quantification

(Designed for RNAscope®)

Indica Labs ISH-IHC measures one IHC stain and up to two chromogenic and/or silver-labeled DNA or RNA ISH probes on a cell-by-cell basis. This allows the user to rapidly contextualize the corresponding protein and gene expression profile of every cell across the tissue. The HALO ISH-IHC module is designed to work with up to 5 stains, including a nuclear stain and up to 4 IHC biomarkers or ISH probes and supports the H-score protocol for RNAscope as recommended by the manufacturer, ACD, a Bio-techne brand.

The module reports numerous outputs including, but not limited to: number of IHC positive cells in total and by cell compartment, Average IHC OD and average IHC OD by cell compartment, percentage IHC positive total and by cell compartment, number and percentage of probe 1 and/or probe 2 positive cells, probe 1 and probe 2 copies per cell and area per cell, cell score for probe 1 and probe 2 (0, +1, +2, +3 and +4), total H-score for each probe, total cells and percentage of total cells positive for IHC marker and probe 1 and/or probe 2 (dual and triple positive cells).   Histograms are automatically generated representing the frequency distribution of probes per cell [Figure 1] and RNAscope cell scoring [Figure 2] as shown below.

Figure 1 shows a distribution curve of probe number on a cell by cell basis, in this example most cells were shown to have between 0-5 copies per cell.

Figure 2. This bar chart shows the numbers of cells that were categorized as 0, 1+, 2+, 3+ and 4+ respectively. The chart shows that a slim majority of cells were positive for RNA probe expression.

The ISH-IHC quantification module integrates seamlessly into the HALO® platform which is compatible with a number of third party systems and  file formats.

File formats supported by the HALO image analysis platform:

  • Non-proprietary (JPG, TIF, OME.TIFF)
  • Nikon (ND2)
  • 3D Histech (MRXS)
  • Akoya (QPTIFF, component TIFF)
  • Olympus / Evident (VSI)
  • Hamamatsu (NDPI, NDPIS)
  • Aperio (SVS, AFI)
  • Zeiss (CZI)
  • Leica (SCN, LIF)
  • Ventana (BIF)
  • Philips (iSyntax, i2Syntax)
  • KFBIO (KFB, KFBF)
  • DICOM (DCM*)
    *whole-slide images

Quantitative RNAscope Image Analysis Guide​

Advance your RNAscope image analysis by downloading the latest Quantitative RNAscope Image Analysis Guide from Indica Labs and ACD, a Bio-techne brand.

RNAscope® Publications with the HALO® Image Analysis Platform

Read this publications list to see examples of research in areas from neuroscience to oncology using HALO to analyze RNAscope images.

Publication Spotlight

The table below includes publications that cite the ISH-IHC and FISH-IF modules. 

Your publication not on the list?  Drop us an email to let us know about it!

TitleAuthorsYearJournalApplicationHALO ModulesProduct
Detection of EpsteinñBarr Virus in Periodontitis: A Review of Methodological ApproachesTonoyan L, Chevalier M, Vincent-Bungnas S, Marsault R, Doglio A2021MircroorganismsReviewISH/FISH, ISH-IHC/FISH-IFHALO
Detection of engineered T cells in FFPE tissue by multiplex in situ hybridization and immunohistochemistryWright JH, Huang L-Y, Weaver S, Archila LD, McAfee MS, Hirayama AV, Chapuis AG, Bleakley M, Rongvaux A, Turtle CJ, Chanthaphavong S, Campbell JS, Pierce RH2020Journal of Immunological MethodsImmunologyISH/FISH, ISH-IHC/FISH-IFHALO
Epicardium-derived cells organize through tight junctions to replenish cardiac muscle in salamandersEroglu E, Yen C, Witman N, Elewa A, Araus A, Wang H, Szattler T, Umeano C, Sohlmer J, Goedel A, Simon A, Chien K2022Nature Cell BiologyMyologyISH/FISH, ISH-IHC/FISH-IFHALO
Satellite repeat RNA expression in epithelial ovarian cancer associates with a tumor immunosuppressive phenotypePorter R, Sun S, Flores M, Berzolla E, You E, Phillips I, KC N, Desai N, Tai E, Szabolcs A, Lang E, Pankaj A, Raabe M, Thapar V, Xu K, Nieman L, Rabe D, Kolin D, Stover E, Pepin D, Stott S, Deshpande V, Liu J, Solovyov A, Matulonis U, Greenbaum B, Ting D2022The Journal of Clinical InvestigationImmuno-oncologyISH/FISH, ISH-IHC/FISH-IFHALO
Systemic Nos2 Depletion and Cox inhibition limits TNBC disease progression and alters lymphoid cell spatial orientation and densitySomasundaram V, Ridnour L, Cheng R, Walke A, Kedei N, Bhattacharyya D, Wink A, Edmondson E, Butcher D, Warner A, Dorsey T, Scheiblin D, Heinz W, Bryant R, Kinders R, Lipkowitz S, Wong S, Pore M, Hewitt S, McVicar D, Anderson S, Chang J, Glynn S, Ambs S, Lockett S, Wink D2022Redox BiologyOncologyClassifer, ISH/FISH, Spatial Analysis, Registration, ISH-IHC/FISH-IFHALO
Annexin A2/TLR2/MYD88 pathway induces arginase 1 expression in tumor-associated neutrophilsZhang H, Zhu X, Friesen T, Kwak J, Pisarenko T, Mekvanich S, Velasco M, Randolph T, Kargl J, Houghton A2022Journal of Clinical InvestigationImmuno-oncologySpatial Analysis, ISH-IHC/FISH-IFHALO
Spatiotemporal transcriptome analysis reveals critical roles for mechano-sensing genes at the border zone in remodeling after myocardial infarctionYamada S, Ko T, Hatsuse S, Nomura S, Zhang B, Dai Z, Inoue S, Kubota M, Sawami K, Yamada T, Sassa T, Katagiri M, Fujita K, Katoh M, Ito M, Harada M, Toko H, Takeda N, Morita H, Aburatani H, Komuro I2022Nature Cardiovascular ResearchMyologyISH-IHC/FISH-IFHALO
Loss of functional System x-c uncouples aberrant postnatal neurogenesis from epileptogenesis in the hippocampus of Kcna1-KO miceAloi M, Thompson S, Quartapella N, Noebels J2022Cell ReportsNeuroscienceISH-IHC/FISH-IFHALO
Multiparameter immunohistochemistry analysis of HIV DNA, RNA and immune checkpoints in lymph node tissueRichardson Z, Deleage C, Tutuka C, Walkiewica M, Del Rio-Estrada P, Pascoe R, Evans V, Reyesteran G, Gonzales M, Roberts-Thomson S, Gonzalez-Navarro M, Torres-Ruiz, F, Estes J, Lewin S, Cameron P2021Journal of Immunological MethodsInfectious DiseaseISH-IHC/FISH-IFHALO
The potassium channel auxiliary subunit Kv_2 (Kcnab2) regulates Kv1 channels and dopamine neuron firingYee J, Rastani A, Soden M2022Journal of NeurophysiologyNeuroscienceISH-IHC/FISH-IFHALO
Transcriptional vulnerabilities of striatal neurons in human and rodent models of Huntington’s diseaseMatsushima A, Pineda S, Crittenden J, Lee H, Galani K, Mantero J, Tombaugh G, Kellis M, Heiman M, Graybiel A2023Nature CommunicationsNeuroscienceArea Quantification, ISH-IHC/FISH-IFHALO
Exploration of spatial heterogeneity of tumor microenvironment in nasopharyngeal carcinoma via transcriptional digital spatial profilingWang L, Wang D, Zeng X, Zhang Q, Wu H, Liu J, Wang Y, Liu G, Poan Y2023International Journal of Biological SciencesOncologyArea Quantification, ISH-IHC/FISH-IFHALO

Related HALO Modules

Tissue Classifier Add-On

Separate multiple tissue classes across a tissue using a learn-by-example approach. Can be used in conjunction with all other modules (fluorescent and brightfield) to select specific tissue classes for further analysis.

Learn More
ISH (RNAscope™)

Quantify spot number, optical density and co-expression of one or two brightfield RNA or DNA probes on a per cell basis. Includes support for brightfield RNAscope assays.

Learn More
FISH (RNAscope™)

Quantify number, intensity and co-expression of an unlimited number of fluorescently labeled RNA or DNA probes on a per cell basis. Includes support for fluorescent RNAscope assays.

Learn More

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