We observed the distribution of lncRNA in the HepG2 by fluorescence in situ hybridization (FISH) Kit (RiboTM lncRNA FISH Probe Mix and RiboTM Fluorescent In Situ Hybridization; RiboBio, China). We used RiboBio to design and synthesize Cy3-labeled LINC317.5 probes, U6 probes, and 18 S probes. Both U6 and 18 S were the internal reference, U6 was almost all located in the nucleus while 18 S was almost all located in the cytoplasm.
Ribotm lncrna fish probe mix
Manufactured by RiboBio
Sourced in China
RiboTM lncRNA FISH Probe Mix is a set of fluorescently labeled DNA probes designed for the detection and visualization of long non-coding RNA (lncRNA) molecules in fixed cells or tissues using Fluorescence In Situ Hybridization (FISH) technique. The probe mix enables the specific targeting and localization of lncRNA transcripts within the cellular context.
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Lab products found in correlation
Ribotm fluorescent in situ hybridization kit, by RiboBio (2 mentions)
Fluorescence microscope, by Olympus (2 mentions)
Ribotm fluorescent in situ hybridization, by RiboBio (1 mentions)
Ribotm fish kit, by RiboBio (1 mentions)
A confocal microscope, by Leica camera (1 mentions)
Fluorescence filter sets, by Zeiss (1 mentions)
Fluorescent in situ hybridization kit, by RiboBio (1 mentions)
7 protocols using ribotm lncrna fish probe mix
1
Localization of lncRNA LINC317.5 in HepG2 cells
2
Visualizing CRNDE-h in CD4+ T Cells
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A cy3-labeled CRNDE-h probe was synthesized using a RiboTM lncRNA FISH Probe Mix (RiboBio, China). Human naive CD4+ T cells were treated with CRC cell-derived exosomes for the indicated times (4 and 24 h). Then, the location of CRNDE-h in the T cells was determined through RNA-FISH by using a RiboTM FISH Kit (RiboBio). In brief, the cells were fixed with 4% paraformaldehyde and then incubated with a transparent buffer. Subsequently, the cells were incubated overnight with 20 µL Cy3-labeled CRNDE-h probe (20 µM) at room temperature in the dark. The nucleus of the cells was stained with DAPI. Finally, the cells were observed under a confocal microscope (Leica, Germany).
3
Localization of lncRNA by FISH
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RiboTM lncRNA FISH Probe Mix (lnc11001001, RIBOBIO) and RiboTM Fluorescent in Situ Hybridization Kit (C10910, RIBOBIO) were used for the FISH of lncRNA, thus detecting the distribution of the target lncRNA. The cell slides were placed at the bottom of a 24-well plate and each well was plated with 1 × 105 cells. After the cells had grown to about 80%, the cells were washed with phosphate-buffered saline (PBS) and fixed with 4% paraformaldehyde. Subsequently, the cells were washed again and treated with permeabilization solution, then 200 μL of prehybridization solution was added and the cells were blocked at 37 °C for 30 min. The prehybridization solution was discarded and 100 μL of the hybridization solution containing the lncRNA FISH probe was added for overnight hybridization at 37 °C. Next day, the cells were washed by PBS and stained with DAPI and photographed by fluorescence microscopy, with 18S and U6 as the reference genes.
4
Subcellular Localization of LncRNA
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FISH assays were performed using a RiboTM fluorescent in situ hybridization kit and RiboTM lncRNA FISH probe mix (RiboBio Co., Ltd.) according to the manufacturer’s protocols. Briefly, cells seeded on chamber slides were fixed in 4% formaldehyde for 10 minutes and then washed with PBS. The cells were incubated with 20 μmol/L lncRNA FISH probe mix at 37°C overnight. After washing, the FISH preparations were counterstained with DAPI and observed by confocal microscopy using appropriate fluorescence filter sets (Zeiss, Oberkochen, Germany). A lncRNA probe labelled with Cy3 was designed and synthesized by RiboBio Co., Ltd., and RiboTM U6 and RiboTM 18S were used as the reference controls for subcellular localization of the lncRNA.
5
Subcellular Localization of ZNF667-AS1 in Leukemic Cells
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FISH was utilized to identify the subcellular localization of ZNF667-AS1 in THP-1 and HL-60 cells. The cover slides were placed in 6-well culture plates, and THP-1 and HL-60 cells were seeded into plates according to RiboTM lncRNA FISH probe Mix (Guangzhou RiboBio Co., Ltd., Guangzhou, Guangdong, China). After a 1-day culture, the cell confluence reached around 80%. The samples were then fixed at room temperature with 1 mL 4% paraformaldehyde and treated with protease K (2 μg/mL), glycine, and ethyl phthalide reagent. Afterwards, the cells were pre-hybridized with 250 μL pre-hybridization solution at 42°C for a period of 1 h and hybridized with 250 μL hybridization solution containing probe (300 ng/mL) at 42°C overnight. The nucleus was then stained with phosphate-buffered saline/Tween-diluted 4ʹ,6-diamidino-2-phenylindole (1:800) for 5 min. Finally, cells were sealed with anti-fluorescent quencher. Focus was placed on five distinct areas, and the cells were counted under a fluorescence microscope (Olympus, Tokyo, Japan).
6
Fluorescent In Situ Hybridization for lncRNA
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Fluorescence in situ hybridization assays were performed using the RiboTM Fluorescent in situ Hybridization Kit and the RiboTM lncRNA FISH Probe Mix (Ribo, Guangzhou, China) according to the manufacturer’s protocols. Briefly, the cells were fixed in 4% formaldehyde for 10 min and then washed with PBS. The cells were incubated with 20 μmol/L lncRNA FISH probe mix at 37°C overnight. After washing, the FISH preparations were counterstained with DAPI observed in confocal microscopy for appropriate fluorescence filter sets (Zeiss, Oberkochen, Germany). The lncRNA probe labeled with Cy3 was designed and synthesized by RiboBio Co., Ltd., RiboTM U6 and RiboTM 18S were used as reference controls for the subcellular localization of lncRNA.
7
LncRNA ADAMTS9-AS2 Expression in Mice Cancer
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The Ribo TM LncRNA FISH Probe Mix (RiboBio Co., Ltd., China) was employed to conduct the FISH assay to investigate the expressions and localization of LncRNA ADAMTS9-AS2 in mice cancer tissues based on the instructions provided by the manufacturer. Briefly, the mice tissues were collected and fixed with 4 % paraformaldehyde at room temperature, and susequently treated with protease K, glycine and acetylation reagent and pre-hybridization solution for 1 h at 42 . After that, the LncRNA ADAMTS9-AS2 probes were incubated with the tissues at 42 overnight. Then, the tissues were stained with DAPI for nucleus. Finally, the tissues were observed and photographed under fluorescence microscope (Olympus, Japan).
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