Before Markup

HALO® Module

ISH Quantification

(Designed for RNASCOPE®)

Indica Labs in situ hybridization (ISH) module is recognized as the fastest, most accurate, and user friendly analysis tool for quantifying nucleic acid probes across tissues and cell lines. This module measures chromogenic and/or silver-labelled DNA or RNA ISH probes on a cell-by-cell basis. This allows the user go rapidly contextualize the gene expression profile of every cell across a tissue. HALO ISH analysis is designed to work with up to three ISH probes and supports the H-score protocol for RNAscope as recommended by the manufacturer, ACD, a Bio-techne brand.

The module reports the total number of cells, total number of spots, total area of spots, the average number of spots per cell and per unit area of tissue, the average area of spots per cell and per unit area of tissue, as well as the number and percentage of cells with RNAscope score of 0, 1+, 2+, 3+ and 4+. When two probes are present, the module also reports the ratio of probe 1: probe 2. 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 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)
  • 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 Quantification module. 

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

TitleAuthorsYearJournalApplicationHALO ModulesProduct
EBV+†tumors exploit tumor cell-intrinsic and -extrinsic mechanisms to produce regulatory T cell-recruiting chemokines CCL17 and CCL22Jorapur A, Marshall L, Jacobson S, Xu M, Marubayashi S, Zibinsky M, Hu D, Robles O, Jackson J, Baloche V, Busson P, Wustrow D, Brockstedt D, Talay O, Kassner P, Cutler G2022PLOS PathogensOncologyISH/FISHHALO
AAV9/MFSD8 gene therapy is effective in preclinical models of neuronal ceroid lipofuscinosis type 7 diseaseChen X, Dong T, Hu Y, Shaffo F, Belur N, Mazzulli J, Gray S2022The Journal of Clinical InvestigationNeuroscienceArea Quantification, ISH/FISHHALO
Genome Sequence and Experimental Infection of Calves with Bovine Gammaherpesvirus 4 (BoHV-4)Bauermann F, Falkenberg S, Martins M, Dassanayake R, Neill J, Ridpath J, Silveira S, Palmer M, Buysse A, Mohr A, Flores E, Diel D2022Research SquareInfectious Disease, OtherISH/FISHHALO
Vertical transmission of attaching and invasive E. coli from the dam to neonatal mice predisposes to more severe colitis following exposure to a colitic insult later in lifeBrand M, Proctor A, Hostetter J, Zhou N, Friedberg I, Jergens A, Phillips G, Wannemuehler M2022PLOS ONEOtherISH/FISHHALO
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
TGF?1 Induces Senescence and Attenuated VEGF Production in Retinal PericytesAvarmovic D, Archaimbault S, Kemble A, Gruener S, Lazendic M, Westenskow P2022BiomedicinesOtherISH/FISHHALO
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
Distinct Encoding of Reward and Aversion by Peptidergic BNST Inputs to the VTASoden M, Yee J, Cuevas B, Rastani A, Elum J, Zweifel L2022Frontiers in Neural CircuitsNeuroscienceISH/FISHHALO
Hyperpolarised†13C-MRI identifies the emergence of a glycolytic cell population within intermediate-risk human prostate cancerSushentsev N, McLean M, Warren A, Benjamin A, Brodie C, Frary A, Gill A, Jones J, Kaggie J, Lamb B, Locke M, Miller J, Mills I, Priest A, Robb F, Shah N, Schulte R, Graves M, Gnanapragasam V, Brindle K, Barrett T, Gallacher F2022Nature CommunicationsOncologyISH/FISH, Membrane, Multiplex IHCHALO
Adult re-expression of IRSp53 rescues NMDA receptor function and social behavior in IRSp53-mutant miceNoh Y, Yook C, Kang J, Lee S, Kim Y, Yang E, Kim H, Kim E2022Communications BiologyNeuroscienceISH/FISH, Figure MakerHALO
The NALCN channel regulates metastasis and nonmalignant cell disseminationRahrmann E, Shorthouse D, Jassim A, Hu L, Ortiz M, Mahler-Araujo B, Vogel P, Paez-Ribes M, Fatemi A, Hannon G, Iyer R, Blundon J, Lourenco F, Kay J, Nazarian R, Hall B, Zakharenko S, Winton D, Zhu L, Gilbertson R2022Nature GeneticsOncologyISH/FISH, Multiplex IHCHALO
Relaxin/insulin-like family peptide receptor 4 (Rxfp4) expressing hypothalamic neurons Q8 modulate food intake and preference in miceLewis J, Woodward O, Nuzzaci D, Smith C, Adriaenssens A, Billing L, Brighton C, Phillips B, Tadross J, Kinston S, Ciabatti E, Gottgens B, Tripodi M, Mornigold D, Baker D, Gribble F, Reimann F2022Molecular MetabolismMetabolismISH/FISH, Highplex FLHALO
Regional epithelial cell diversity in the small intestine of pigsWiarda J, Becker S, Sivasankaran S, Loving C, 2022Journal of Animal ScienceGastroenterologyISH/FISH, Figure MakerHALO
Single cell analysis of cribriform prostate cancer reveals cell intrinsic and tumor microenvironmental pathways of aggressive diseaseWong H, Sheng Q, Hesterberg A, Croessmann S, Rios B, Giri K, Jackson J, Miranda A, Watkins E, Schaffer K, Donahue M, Winkler E, Penson D, Smith J, Herrell S, Luckenbaugh A, Barocas D, Kim Y, Graves D, Giannico G, Rathmell J, Park B, Gordetsky J, Hurley P2022Nature CommunicationsOncologyISH/FISHHALO
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
Glutaminase 2 Knockdown Reduces Hyperammonemia and Associated Lethality of Urea Cycle Disorder Mouse ModelMao X, Chen H, Lin A, Kim S, Burczynski M, Na E, Halasz G, Sleeman M, Murphy A, Okamoto H, Cheng X2022Journal of Inherited Metabolic DiseaseMetabolismArea Quantification, ISH/FISHHALO
What Happens to the Preserved Renal Parenchyma After Clamped Partial Nephrectomy?Xiong L, Nguyen J, Peng Y, Zhou Z, Ning K, Jia N, Nie J, Wen D, Wu Z, Roversi G, Palacios D, Abramczyk E, Munoz-Lopez C, Campbell J, Cao Y, Li Wencai, Zhang X, He Z, Li X, Huang J, Shou J, Wu J, Chen M, Chen X, Zheng J, Xu C, Zhong W, Li Z, Dong W, Zhou J, Zhang H, Luo J, Liu J, Sun G, Han H, Guo S, Dong P, Zhou F, Yu C, Campbell S, Zhang Z2022European UrologyOtherArea Quantification, ISH/FISHHALO
Adolescent intermittent ethanol exposure produces Sex-Specific changes in BBB Permeability: A potential role for VEGFAVore A, Barney T, Deak M, Varlinskaya E, Deak T2022Brain, Behavior, and ImmunityNeuroscienceISH/FISHHALO
Relationship between impaired BMP signalling and clinical risk factors at early-stage vascular injury in the preterm infantHeydarian M, Oak P, Zhang X, Kamgari N, Kindt A, Koschlig M, Pritzke T, Gonzalez-Rodriguez E, Forster K, Morty R, Hafner F, Hubener C, Flemmer A, Yildirim A, Sudheendra D, Tian X, Petrera A, Kirsten H, Ahnert P, Morrell N, Desai T, Sucre J, Spiekerkoetter E, Hilgendorff A2022ThoraxOtherISH/FISHHALO
Spatiotemporal expression pattern of Progesterone Receptor Component (PGRMC) 1 in endometrium from patients with or without endometriosis or adenomyosisThieffry C, Van Wynendaele M, Samain L, Tyteca D, Pierreux C, Marbaix E, Henriet P2022The Journal of Steroid Biochemistry and Molecular BiologyOtherISH/FISH, Highplex FLHALO, HALO AI
Quantitative Imaging Analysis of the Spatial Relationship between Antiretrovirals, Reverse Transcriptase Simian-Human Immunodeficiency Virus RNA, and Fibrosis in the Spleens of Nonhuman PrimatesDevanathan A, White N, Desyaterik Y, De la Cruz G, Nekorchuk M, Terry M, Busman-Sahay K, Adamson L, Luciw P, Fedoriw Y, Estes J, Rosen E, Kashuba A2022Antimicrobial Agents and ChemotherapyInfectious DiseaseISH/FISHHALO
CD123-directed allogeneic chimeric-antigen receptor T-cell therapy (CAR-T) in blastic plasmacytoid dendritic cell neoplasm (BPDCN): Clinicopathological insightsPemmaraju N, Wilson N, Senapati J, Economides M, Guzman M, Neelapu S, Kazemimood R, Davis R, Jain N, Khoury J, Sugita M, Cai T, Smith J, Frattini M, Garton A, Roboz G, Konopleva M2022Leukemia ResearchImmuno-oncologyISH/FISHHALO
Localization of macrophage subtypes and neutrophils in the prostate tumor microenvironment and their association with prostate cancer racial disparitiesMaynard J, Godwin T, Lu J, Vidal I, Lotan T, De Marzo A, Joshu C, Sfanos K2022The ProstateOncologyISH/FISH, Multiplex IHCHALO
Molecular identification of spatially distinct anabolic responses to mechanical loading in murine cortical boneChlebek C, Moore J, Ross F, van der Meulen M2022Journal of Bone and Mineral ResearchOtherISH/FISHHALO
Sex-specific effects of ethanol consumption in older Fischer 344 rats on microglial dynamics and A_(1-42) accumulationMarsland P, Vore A, DaPrano E, Paluch J, Blackwell A, Varlinskaya E, Deak T2022AlcoholNeuroscienceISH/FISHHALO
Estrogen receptor beta activity contributes to both tumor necrosis factor alpha expression in the hypothalamic paraventricular nucleus and the resistance to hypertension following angiotensin II in female miceWoods C, Contoreggi N, Johnson M, Milner T, Wang G, Glass M, 2022Neurochemistry InternationalNeuroscienceISH/FISHHALO
Angiotensin II Infusion Results in Both Hypertension and Increased AMPA GluA1 Signaling in Hypothalamic Paraventricular Nucleus of Male but not Female MiceWang G, Woods C, Johnson M, Milner T, Glass M2022NeuroscienceNeuroscienceISH/FISHHALO
Multiregional single-cell dissection of tumor and immune cells reveals stable lock-and-key features in liver cancerMa L, Heinrich S, Wang L, Keggenhoff F, Khatib S, Forgues M, Kelly M, Hewitt S, Saif A, Hernandez J, Mabry D, Kloeckner R, Greten T, Chaisaingmongkol J, Ruchirawat M, Marquardt J, Wang X2022Nature CommunicationsOncologyISH/FISHHALO
Autologous T cell therapy for MAGE-A4+ solid cancers in HLA-A*02+ patients: a phase 1 trialHong D, Van Tine B, Biswas S, McAlpine C, Johnson M, Olszanski A, Clarke J, Araujo D, Blumenschein G, Kebriaei P, Lin Q, Tipping A, Sanderson J, Wang R, Trivedi T, Annareddy T, Bai J, Rafail S, Sun A, Fernandes L, Navenot J-M, Bushman F, Everett J, Karadeniz D, Broad R, Isabelle M, Naidoo R, Bath N, Betts G, Wolchinsky Z, Batrakou D, Van Winkle E, Elefant E, Ghobadi A, Cashen A, Grand'Maison A, McCarthy P, Fracasso P, Norry E, Williams D, Druta M, Liebner D, Odunsi K, Butler M2023Nature MedicineImmuno-oncologyISH/FISH, Spatial Analysis, Highplex FLHALO
Combined KRASG12C and SOS1 inhibition enhances and extends the anti-tumor response in KRASG12C-driven cancers by addressing intrinsic and acquired resistanceThatikonda V, Lu H, Jurado S, Kostyrko K, Bristow C, Bosch K, Feng N, Gao S, Gerlach D, Gmachi M, Lieb S, Jeschko A, Machado A, Marszalek E, Mahendra M, Jaeger P, Sorokin A, Strauss S, Trapani F, Kopetz S, Vellano C, Petronczki M, Kraut N, Heffernan T, Marszalek J, Pearson M, Waizenegger I, Hofmann M2023bioRxivOncologyISH/FISH, Multiplex IHC, Highplex FL, Figure MakerHALO
Prefrontal cortical protease TACE/ADAM17 is involved in neuroinflammation and stress-related eating alterationsSharafeddin F, Ghaly M, Simon T, Ontiveros-Angel P, Figueroa J2023bioRxivNeuroscienceISH/FISHHALO
Long noncoding RNA LINC01594 inhibits the CELF6-mediated splicing of oncogenic CD44 variants to promote colorectal cancer metastasis.Liu B, Song A, Gui P, Wang J, Pan Y, Li C, Li S, Zhang Y, Jiang T, Xu Y, Huo F, Pei D, Song J2023Research SquareOncologyISH/FISHHALO
CD47-Dependent Regulation of Immune Checkpoint Gene Expression and MYCN mRNA Splicing in Murine CD8 and Jurkat T CellsKaur S, Awad D, Finney R, Meyer T, Singh S, Cam M, Karim B, Warner A, Roberts D2023International Journal of Molecular SciencesImmuno-oncologyISH/FISHHALO
SARS-CoV-2 infection induces DNA damage, through CHK1 degradation and impaired 53BP1 recruitment, and cellular senescenceGioia U, Tavella S, Martinez-Orellana P, Cicio G, Colliva A, Ceccon M, Cabrini M, Henriques A, Fumagalli V, Paldino A, Presot E, Rajasekharan S, Iacomino N, Pisati F, Matti V, Sepe S, Conte M, Barozzi S, Lavagnino Z, Carletti T, Volpe M, Cavalcante P, Iannacone M, Rampazzo C, Bussani R, Tripodo C, Zacchigna S, Marcello A, di Fagagna F2023Nature Cell BiologyInfectious DiseaseISH/FISH, Highplex FLHALO
A cellular and molecular spatial atlas of dystrophic muscleStec M, Su Q, Adler C, Zhang L, Golann D, Khan N, Panagis L, Villalta S, Ni M, Wei Y, Walls J, Murphy A, Yancopoulos G, Atwal G, Kleiner S, Halasz G, Sleeman M2023Proceedings of the National Academy of SciencesMyologyISH/FISH, Highplex FLHALO

Related HALO Modules

TMA (Tissue Microarray) Add-On

Module to assist in the segmentation of TMA spots for further analysis.

Learn More
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
Multiplex 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

Use the arrows above to view additional related modules

Want to Learn More?

Fill out the form below to request information about any of our software products.
You can also drop us an email at