In situ hybridization was performed with an RNAscope Multiplex Fluorescent Detection Reagents v2 kit (Advanced Cell Diagnostics 323110) using the following probes: Col10a1 (426181), Runx2 (414021), Pth1r (426191), Ihh (413091), and Sp7 (403401). After RNAscope, sections were incubated in blocking buffer (3% bovine serum albumin/Tris-buffered saline with Tween (TBST)) for 30 min at room temperature (RT), and subsequently stained with SOX9 polyclonal antibody (1:500; R&D systems, AF3075), RUNX2 polyclonal antibody (1:100; Novus Biological, NBP1-89104), PTHrP polyclonal antibody (1:100; Invitrogen, PA5-57493), PTH1R monoclonal antibody (3D1.1) (1:200; Novus Biological, NBP1-40067), DsRed Polyclonal Antibody (Living Colors) (1:700; Takara Bio, 632496), or mCherry monoclonal antibody (16D7) (1:400; Invitrogen, M11217), overnight at 4°C. Sections were subsequently stained with appropriate Alexa Fluor–conjugated secondary antibodies for 3 h at 4°C, followed by DAPI (4′,6-diamidino-2-phenylindole, 5 µg/mL; Invitrogen D1306) staining. More detailed experimental procedures are available in the Appendix Materials and Methods .
Rnascope multiplex fluorescent detection reagents v2 kit
Manufactured by Advanced Cell Diagnostics
The RNAscope Multiplex Fluorescent Detection Reagents v2 kit is a laboratory equipment product designed for the detection and visualization of RNA targets in fixed cells and tissues. The kit provides the necessary reagents and tools for performing multiplex fluorescent in situ hybridization (FISH) experiments, allowing the simultaneous detection of up to four different RNA targets within a single sample.
Automatically generated - may contain errors
Lab products found in correlation
Nbp1 89104, by Thermo Fisher Scientific (1 mentions)
D1306, by Thermo Fisher Scientific (1 mentions)
Opal 570 fp1488001kt, by PerkinElmer (1 mentions)
Lsm 800, by Zeiss (1 mentions)
Rnascope probe, by Advanced Cell Diagnostics (1 mentions)
Alexa fluor 546 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Living colors dsred polyclonal antibody, by Takara Bio (1 mentions)
4 protocols using rnascope multiplex fluorescent detection reagents v2 kit
1
Spatial Gene Expression Analysis of Cartilage Development
2
Grin1 mRNA Expression in SMACreER:Ai14 Mice
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According to the manufacturer’s protocol, the RNAscope assay was performed using the RNAscope Multiplex Fluorescent Detection Reagents v2 Kit (323110, Advanced Cell Diagnostics). Briefly, frozen brain sections from adult SMACreER:Ai14 were baked at 60 °C for 1 h, rinsed with PBS, and treated with hydrogen peroxide at room temperature for 10 min. The target retrieval was performed at 98–102 °C for 5 min, followed by protease plus (322331) treatment at 42 °C for 30 min. The Grin1 RNAscope probe (431611) was then hybridized at 42 °C for 2 h. Then, the slices were subjected to RNAscope amplification and chromogenic detection (Opal 570, FP1488001KT, PerkinElmer). The slices were stained with DAPI, mounted with an antifade mounting medium, and imaged with a fluorescence microscope (Carl Zeiss LSM 800) acquired by Zeiss ZEN software (blue edition, version 3.6).
3
SARS-CoV-2 Detection in Fungiform Papillae
4
RNAscope Multiplex Fluorescent Detection
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In situ hybridization was performed with RNAscope Multiplex Fluorescent Detection Reagents v2 kit (Advanced Cell Diagnostics, catalog 323110) using the following riboprobes: Ptch1exon8–9 (specific to exons 8 and 9 flanked by loxP sites, catalog 476231) and Smo (catalog 318411). Briefly, sections were treated with H2O2 for 10 minutes and dehydrated with 100% ethanol for 3 minutes at room temperature. Samples were subsequently treated with RNAscope Protease III for 30 minutes at 40°C and washed with distilled water. Sections were treated with each target probe and hybridized for 2 hours, at 40°C, followed by signal hybridization by AMP and amplification by HRP-C1. TSA Vivid Fluorophore 650 (1:1,000, Advanced Cell Diagnostics, 323273) was added to label the C1 probe, and sections were treated with HRP blocker. After washing with wash buffer, sections were incubated in blocking buffer consisting of 3% bovine serum albumin/TBST for 30 minutes, at room temperature, and stained with Living Colors DsRed Polyclonal Antibody (1:800, Takara Bio, 632496) overnight at 4°C. Sections were subsequently stained with Alexa Fluor 546–conjugated goat anti-rabbit IgG (A11035, 1:400, Invitrogen) for 3 hours, at 4°C, followed by DAPI staining. Signals from RNAscope assays were quantified by using Zeiss Zen software (v3.0).
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