ACD can configure probes for the various manual and automated assays for INS for RNAscope Assay, or for Basescope Assay compatible for your species of interest.
Applied In Vitro Toxicology
2019 Feb 27
Neau L, Lorin C, Frentzel S, Hoeng J, Iskandar A, Leroy P, Trivedi K.
PMID: - | DOI: 10.1089/aivt.2018.0021
Abstract
Introduction: Developed by Advanced Cell Diagnostics, RNAscope™in situ hybridization technology enables detection of a target RNA in a cell-specific manner on formalin-fixed paraffin-embedded tissue sections and represents a good alternative to immunohistochemistry. The goal of this work is to illustrate an optimized protocol of the RNAscope technology to detect target genes in various human organotypic culture models (nasal, small airway, and gingival). These culture models retain the three-dimensional structure of native epithelium, mimic in vivo morphology and human physiology, and can be used as alternative sources to animal testing.
Materials and Methods: After fixation and processing of five replicates of the three different organotypic cell cultures, the tissue morphology was checked by hematoxylin and eosin staining. The RNAscope protocols were optimized based on three crucial parameters: heat pretreatment, enzymatic digestion, and signal amplification. Digital images of the RNAscope stained slides were generated using the Hamamatsu NanoZoomer 2.0 slide scanner, and images were quantified using a custom-made plugin on Definiens Tissue Studio software (Definiens AG, Munich, Germany).
Results: The tissue morphology demonstrates optimum fixation and processing for samples, while the optimized protocol for RNAscope shows preserved RNA with staining on the positive control probe with score ≥2 and no staining on the negative control probe with score <1.
Discussion and Conclusion: RNAscope combined with organotypic cell cultures is a promising tool to better understand cell-specific RNA expression while implementing 3R (replace, reduce, and refine animal testing) principles
bioRxiv : the preprint server for biology
2023 Feb 05
Su, Y;Xu, J;Zhu, Z;Yu, H;Nudell, V;Dash, B;Moya, EA;Ye, L;Nimmerjahn, A;Sun, X;
PMID: 36778350 | DOI: 10.1101/2023.02.04.527145
Kidney international
2022 May 26
Kaneko, K;Sato, Y;Uchino, E;Toriu, N;Shigeta, M;Kiyonari, H;Endo, S;Fukuma, S;Yanagita, M;
PMID: 35644281 | DOI: 10.1016/j.kint.2022.04.026
Bio Protoc.
2017 Nov 20
Fe Lanfranco M, Loane DJ, Mocchetti I, Burns MP, Villapol S.
PMID: 29238736 | DOI: 10.21769/BioProtoc.2608
Microglia and macrophage cells are the primary producers of cytokines in response to neuroinflammatory processes. But these cytokines are also produced by other glial cells, endothelial cells, and neurons. It is essential to identify the cells that produce these cytokines to target their different levels of activation. We used dual RNAscope® fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) techniques to visualize the mRNA expression pattern of pro- and anti-inflammatory cytokines in microglia/macrophages cells. Using these methods, we can associate one mRNA to specific cell types when combining with different cellular markers by immunofluorescence. Results from RNAscope® probes IL-1β, TNFα, TGFβ, IL-10 or Arg1, showed colocalization with antibodies for microglia/macrophage cells. These target probes showed adequate sensitivity and specificity to detect mRNA expression. New FISH detection techniques combined with immunohistochemical techniques will help to jointly determine the protein and mRNA localization, as well as provide reliable quantification of the mRNA expression levels.
Cell Rep.
2018 Jan 09
Chevée M, Robertson JJ, Cannon GH, Brown SP, Goff LA.
PMID: 29320739 | DOI: 10.1016/j.celrep.2017.12.046
Single-cell RNA sequencing has generated catalogs of transcriptionally defined neuronal subtypes of the brain. However, the cellular processes that contribute to neuronal subtype specification and transcriptional heterogeneity remain unclear. By comparing the gene expression profiles of single layer 6 corticothalamic neurons in somatosensory cortex, we show that transcriptional subtypes primarily reflect axonal projection pattern, laminar position within the cortex, and neuronal activity state. Pseudotemporal ordering of 1,023 cellular responses to sensory manipulation demonstrates that changes in expression of activity-induced genes both reinforced cell-type identity and contributed to increased transcriptional heterogeneity within each cell type. This is due to cell-type biased choices of transcriptional states following manipulation of neuronal activity. These results reveal that axonal projection pattern, laminar position, and activity state define significant axes of variation that contribute both to the transcriptional identity of individual neurons and to the transcriptional heterogeneity within each neuronal subtype.
Neuron.
2017 Jan 31
François A, Low SA, Sypek EI, Christensen AJ, Sotoudeh C, Beier KT, Ramakrishnan C, Ritola KD, Sharif-Naeini R, Deisseroth K, Delp SL, Malenka RC, Luo L, Hantman AW, Scherrer G.
PMID: 28162807 | DOI: 10.1016/j.neuron.2017.01.008
Pain thresholds are, in part, set as a function of emotional and internal states by descending modulation of nociceptive transmission in the spinal cord. Neurons of the rostral ventromedial medulla (RVM) are thought to critically contribute to this process; however, the neural circuits and synaptic mechanisms by which distinct populations of RVM neurons facilitate or diminish pain remain elusive. Here we used in vivo opto/chemogenetic manipulations and trans-synaptic tracing of genetically identified dorsal horn and RVM neurons to uncover an RVM-spinal cord-primary afferent circuit controlling pain thresholds. Unexpectedly, we found that RVM GABAergic neurons facilitate mechanical pain by inhibiting dorsal horn enkephalinergic/GABAergic interneurons. We further demonstrate that these interneurons gate sensory inputs and control pain through temporally coordinated enkephalin- and GABA-mediated presynaptic inhibition of somatosensory neurons. Our results uncover a descending disynaptic inhibitory circuit that facilitates mechanical pain, is engaged during stress, and could be targeted to establish higher pain thresholds.
JCI Insight
2020 Feb 27
Celik M�, Labuz D, Keye J, Glauben R, Machelska H
PMID: 32102987 | DOI: 10.1172/jci.insight.133093
Neuron
2022 Oct 26
Albisetti, GW;Ganley, RP;Pietrafesa, F;Werynska, K;Magalhaes de Sousa, M;Sipione, R;Scheurer, L;Bösl, MR;Pelczar, P;Wildner, H;Zeilhofer, HU;
PMID: 36323322 | DOI: 10.1016/j.neuron.2022.10.008
Cell reports
2022 May 31
Mu, R;Tang, S;Han, X;Wang, H;Yuan, D;Zhao, J;Long, Y;Hong, H;
PMID: 35649349 | DOI: 10.1016/j.celrep.2022.110882
EMBO reports
2021 Sep 27
Chen, W;Liu, X;Li, W;Shen, H;Zeng, Z;Yin, K;Priest, JR;Zhou, Z;
PMID: 34569705 | DOI: 10.15252/embr.202152389
Nature
2022 Mar 01
Zhao, Q;Yu, CD;Wang, R;Xu, QJ;Dai Pra, R;Zhang, L;Chang, RB;
PMID: 35296859 | DOI: 10.3760/cma.j.cn112151-20210719-00516
Cell reports
2022 Dec 13
Huang, XT;Li, T;Li, T;Xing, S;Tian, JZ;Ding, YF;Cai, SL;Yang, YS;Wood, C;Yang, JS;Yang, WJ;
PMID: 36516755 | DOI: 10.1016/j.celrep.2022.111796
Description | ||
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sense Example: Hs-LAG3-sense | Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe. | |
Intron# Example: Mm-Htt-intron2 | Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection | |
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G) | A mixture of multiple probe sets targeting multiple genes or transcripts | |
No-XSp Example: Hs-PDGFB-No-XMm | Does not cross detect with the species (Sp) | |
XSp Example: Rn-Pde9a-XMm | designed to cross detect with the species (Sp) | |
O# Example: Mm-Islr-O1 | Alternative design targeting different regions of the same transcript or isoforms | |
CDS Example: Hs-SLC31A-CDS | Probe targets the protein-coding sequence only | |
EnEm | Probe targets exons n and m | |
En-Em | Probe targets region from exon n to exon m | |
Retired Nomenclature | ||
tvn Example: Hs-LEPR-tv1 | Designed to target transcript variant n | |
ORF Example: Hs-ACVRL1-ORF | Probe targets open reading frame | |
UTR Example: Hs-HTT-UTR-C3 | Probe targets the untranslated region (non-protein-coding region) only | |
5UTR Example: Hs-GNRHR-5UTR | Probe targets the 5' untranslated region only | |
3UTR Example: Rn-Npy1r-3UTR | Probe targets the 3' untranslated region only | |
Pan Example: Pool | A mixture of multiple probe sets targeting multiple genes or transcripts |
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