Strataquest software

Manufactured by TissueGnostics

Sourced in Austria

StrataQuest is a software solution designed for image analysis and quantification. It provides a comprehensive platform for processing, visualizing, and analyzing digital images generated from various imaging modalities. The software offers a range of tools and functionalities to support researchers in their data analysis and decision-making processes.

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StrataQuest (26 citations) StrataQuest analysis software (10 citations) StrataQuest image analysis software (2 citations) StrataQuest tissue analysis software (2 citations)

27 protocols using strataquest software

Quantitative IHC Analysis of NEK2 in ccRCC

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Immunohistochemical (IHC) analysis was conducted using tissue microarray (TMA) of ccRCC and adjacent normal tissue specimens according to the protocol of the Two-Step IHC Kit (ZSGB-BIO, Beijing, China) using an anti-NEK2 antibody (ab227958, 1:200; Abcam, Cambridge, UK). The specimens of TMA were reviewed by one uropathologist and the most representative areas of renal tumor cells and adjacent tumor stroma were selected for the donor block. The TMAs were made by using a tissue-arraying instrument (Manual Tissue Microarrayer Quick-Ray ®). Cores with 2 mm diameter from per-donor block were diverted into a recipient block microarray.
The StrataQuest software (TissueGnostics, Vienna, Austria) was used to scan and analyze the images to quantify the tumor tissues, para-carcinoma tissues and NEK2 positive cells in the TMA. We multiplied the quantitative value of positive staining intensity (negative = 1, weak = 2, moderate = 3, strong = 4) by percentages of positive staining (0% = 0, ≤25% = 1, 26%–50% = 2, 51%–75% = 3, ≥76% = 4), and the data obtained was weighted and set into four levels (Ⅰ, Ⅱ, Ⅲ, Ⅳ). All results were verified by three experienced independent investigators.

Wang C., Huang Y., Ma X., Wang B, & Zhang X. (2021). Overexpression of NEK2 is correlated with poor prognosis in human clear cell renal cell carcinoma. International Journal of Immunopathology and Pharmacology, 35, 20587384211065893.

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Quantitative Multiplex Fluorescence Imaging

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Using the corresponding phase-contrast images, fluorescence images produced after each antibody stain were aligned pixelwise and were corrected for illumination faults using flat-field correction. The alignment reached a resolution of ±1 pixel. Post-bleaching images were subtracted from the following fluorescence tag images. Superimposed images composed a n epitope expression in relation to each pixel (900 × 900-nm² area) of a visual field (1,024 × 1,024 pixels).
Protein expression quantification was conducted by StrataQuest software (TissueGnostics) and is described in Fig S5.

Schierer S., Ostalecki C., Zinser E., Lamprecht R., Plosnita B., Stich L., Dörrie J., Lutz M.B., Schuler G, & Baur A.S. (2018). Extracellular vesicles from mature dendritic cells (DC) differentiate monocytes into immature DC. Life Science Alliance, 1(6), e201800093.

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Quantitative Immunofluorescence Analysis of Pancreatic Tissue

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IF was conducted using FFPE pancreatic tissue sections and performed as described in detail previously^{5 (link),6 (link),30 (link),31 (link),37 (link)}. All IF images were acquired using the TissueFAXS system and further analyzed by the StrataQuest software (TissueGnostics, Vienna, Austria), allowing the quantification of target-protein positive cell numbers from DAPI positive cells or amylase-specific positive cells (indicating pancreatic acinar cells), as previously described in detail^{5 (link),6 (link),31 (link)}. Infiltrated MPO-positive monocytes/ neutrophils were determined by the numbers of positive cell per mm² tissue. All other chemicals were from Sigma-Aldrich (St. Louis, USA) or ROTH (Karlsruhe, Germany), if not stated otherwise.

Xia L., Xu Z., Zhou X., Bergmann F., Grabe N., Büchler M.W., Neoptolemos J.P., Hackert T., Kroemer G, & Fortunato F. (2020). Impaired autophagy increases susceptibility to endotoxin-induced chronic pancreatitis. Cell Death & Disease, 11(10), 889.

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Multiplex Immunohistochemistry Analysis of Glioma Tissue

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We purchased the glioma tissue array from Wuhan Tanda Scientific Co., Ltd. (NGL1021), with ethics approvement. SOX10 (Mouse, 1:100, Proteintech, China), CD163 (Rabbit, 1:3,000, Proteintech, China), and CD68 (Rabbit, 1:3,000, Servicebio, China) were the primary Abs. Horseradish peroxidase-conjugated secondary antibody incubation (GB23301, GB23303, Servicebio, China) was the secondary antibody. The tyramide signal was amplified into TSA [FITC-TSA, CY3-TSA, 594-TSA, and 647-TSA (Servicebio, China)]. The stained slides were scanned using the TissueFAXS platform (TissueGnostics, Vienna, Austria). The spatial analysis of the stained cells was performed using the StrataQuest software (TissueGnostics, Vienna, Austria).

Xiao G., Wang K., Wang Z., Dai Z., Liang X., Ye W., Luo P., Zhang J., Liu Z., Cheng Q, & Peng R. (2022). Machine learning-based identification of SOX10 as an immune regulator of macrophage in gliomas. Frontiers in Immunology, 13, 1007461.

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Histological Analysis of Hippocampus

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Brain tissue was fixed in 4% paraformaldehyde, dehydrated in increasing concentrations of alcohol, and embedded in paraffin. Specimens were cut into 4 μm sections using a section cutter and stained with hematoxylin–eosin (H&E, Servicebio Technology Co., Ltd., Wuhan, China) and Nissl stain (Servicebio Technology Co., Ltd., Wuhan, China) according to the manufacturer’s instructions. The number of cells in the hippocampus was quantitated using the StrataQuest software (version 7.0.1, TissueGnostics GmbH, Vienna, Austria).

Guo P., Zeng M., Wang S., Cao B., Liu M., Zhang Y., Jia J., Zhang Q., Zhang B., Wang R., Zheng X, & Feng W. (2022). Eriodictyol and Homoeriodictyol Improve Memory Impairment in Aβ25–35-Induced Mice by Inhibiting the NLRP3 Inflammasome. Molecules, 27(8), 2488.

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Cell Adhesion and Migration Assay on PEDOT-co-PDLLA Scaffolds

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Cell
adhesion was evaluated by the number of EBs able to attach onto the
polymeric substrates. After formation and neural induction, 30 EBs
were plated onto traditional plastic surfaces or PEDOT-co-PDLLA-based scaffolds with increasing concentrations of PEDOT. Cells
were incubated at ideal conditions for 24 h. Then, the medium with
unattached EBs was changed. EBs adhered onto the polymeric surfaces
were counted. To measure cell migration from the EBs, images were
recorded after 48 h, and migration halos were analyzed and quantified
with StrataQuest software (TissueGnostics). This imaging analysis
program analyzed data obtained from tissue or cell culture slides
using a workflow similar to that of flow cytometry. Gating procedures
were used to determine cell populations positive or negative for epitope
expression of interest. DAPI fluorescence was used as a master channel
for cell quantification. Images of phase-contrast and DAPI and MAP2-staining
Alexa488 fluorescence channels were overlapped. Migration halo analysis
was performed by combining and processing the images of DAPI and phase-contrast
ROI. Total length measurements of anti-MAP2-tagged neurites were performed.

da Silva A.C., Semeano A.T., Dourado A.H., Ulrich H, & Cordoba de Torresi S.I. (2018). Novel Conducting and Biodegradable Copolymers with Noncytotoxic Properties toward Embryonic Stem Cells. ACS Omega, 3(5), 5593-5604.

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Beiging Regulation by Adipose ERα

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To determine the role of adipose tissue ERα to mediate beiging, ERα was added back to iWAT of female ERKO mice. Briefly, C57BL6 WT and ERKO female mice, housed in normal vivarium conditions (22 °C), were injected with an ERα overexpression vector (Ad-ERα) in one iWAT fat pad and a control vector (Ad-GFP) in the contra-lateral iWAT using the methods previously reported [26] (link). iWAT was collected 6 days after injections and samples were fixed in 10% neutral buffered formalin for 48 h, then embedded in paraffin wax for histology performed by the Cedars-Sinai Biobank and Translational Medicine Core (Los Angeles, CA). Slide images were acquired using Apeiro AT Turbo (Leica Biosystems Inc., U.S.). The average fluorescence intensity of 10 randomly selected iWAT regions of interest (ROI) were measured in each sample using StrataQuest software (TissueGnostics, Austria). Results are expressed as arbitrary units comparing the Ad-ERα iWAT to the contralateral Ad-GFP fat pad.

Santos R.S., Frank A.P., Fátima L.A., Palmer B.F., Öz O.K, & Clegg D.J. (2018). Activation of estrogen receptor alpha induces beiging of adipocytes. Molecular Metabolism, 18, 51-59.

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Automated Pathological Image Analysis

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High-quality images were obtained with TissueFAXS 200 digital pathological analysis system. Data analysis was performed using StrataQuest software (TissueGnostics) [12 (link)]. The software programs are based on single cell detection by identification of nuclear structures [13 ,14 ]. Nuclei and neutrophils were identified using the “disassembly” function. Furthermore, the average nuclear size, discrimination area, discrimination gray, and background threshold for the master marker was specified. All images were analyzed with the same settings after adjustments. Forward and backward gating was routinely used for quality control. Backward gating was used to verify data by visual inspection on the original image. The results are visualized on dot plot scatter grams and/or histograms. Cut-offs (to differentiate between neutrophils and no-neutrophils) and gates (nuclei and no-nuclei) were set in the scatter grams.

Niu J., An G., Gu Z., Li P., Liu Q., Bai R., Sun J, & Du Q. (2020). Analysis of sensitivity and specificity: precise recognition of neutrophils during regeneration of contused skeletal muscle in rats. Forensic Sciences Research, 7(2), 228-237.

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Multiparametric Image Quantification and Analysis

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Image processing and analysis was performed using StrataQuest software version 6.0.1.145 (TissueGnostics, Vienna, Austria). Image processing included reconstruction of whole images and creation of in silico multiplexed images. Image analysis included cell phenotyping and tissue cytometry. A proprietary image stitching algorithm was performed on the acquired tiled images to reconstruct the whole image. Algorithms were created to align the stitched images, generate a composite image containing all biomarkers from all staining rounds, isolate each cell in the composite image, and identify the positive cells in the composite image. Tissue cytometry and backgating into the tissue images were used for quantitation and visualization of the in silico data. The nonproprietary details are as follows:

Blenman K.R, & Bosenberg M.W. (2018). Immune cell and cell cluster phenotyping, quantitation, and visualization using in silico multiplexed images and tissue cytometry. Cytometry. Part A : the journal of the International Society for Analytical Cytology, 95(4), 399-410.

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Wound Healing Assay in mESCs

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Mouse embryonic stem cells (E14Tg2A) were plated on gelatin-coated (0.1% porcine skin gelatin, Sigma-Aldrich) plastic dishes (24 wells plate, Sarstedt). With a p20 pipette tip, a straight scratch was designed when the confluence had reached 70% (24 hs). Then, the culture medium was changed in the presence of the adenosine receptor antagonists. The cultures were scanned using the TissueFAXS Q cytometer (TissueGnostics) every 12 h for 3 days. The areas of the scratch were automatically measured with the StrataQuest software (TissueGnostics). The variation of the area was calculated by subtracting the area of the last picture from the area of the first one (Δ area).

Glaser T., Martins P., Beco R., Bento C.A., Cappellari A.R., La Banca Oliveira S., Merkel C.A., Arnaud-Sampaio V.F., Lameu C., Battastini A.M, & Ulrich H. (2024). Impairment of adenosine signaling disrupts early embryo development: unveiling the underlying mechanisms. Frontiers in Pharmacology, 14, 1328398.

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