FISH was performed using the lncRNA FISH Kit (Guangzhou RiboBio, China) according to the manufacturer's instructions. Briefly, cells were fixed with 4% formaldehyde for 10 min at room temperature. After washing, cells were permeabilized with 0.5% Triton X‐100 for 30 min at 37°C. Next, cells were incubated with RNA probes in hybridization buffer overnight at 37°C. The RNA probes were directly conjugated with a fluorophore. Then, the cells were washed three times with saline sodium citrate buffer, stained with 4′,6‐diamidino‐2‐phenylindole.
Lncrna fish kit
Manufactured by RiboBio
Sourced in China
The LncRNA FISH Kit is a laboratory equipment product designed for the detection and visualization of long non-coding RNA (lncRNA) molecules within cells. It provides the necessary reagents and protocols for performing fluorescence in situ hybridization (FISH) to localize and analyze the expression patterns of specific lncRNA targets.
Automatically generated - may contain errors
Lab products found in correlation
Fish probe, by RiboBio (3 mentions)
Fv1000 confocal microscope, by Olympus (3 mentions)
Lsm 710 confocal microscope, by Zeiss (2 mentions)
Ab181861, by Abcam (1 mentions)
Lsm800 confocal microscope, by Zeiss (1 mentions)
Lsm 700 meta, by Zeiss (1 mentions)
Alexa fluor 488 conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
Nbp1 04676, by Novus Biologicals (1 mentions)
Fish kit, by RiboBio (1 mentions)
12 protocols using lncrna fish kit
1
In Situ Hybridization of lncRNA
2
FISH Analysis of lncRNA TCONS_00323213 in PSCs
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A lncRNA FISH kit (RiboBio, Guangzhou, China) was used for an RNA FISH analysis of PSCs in accordance with the operating procedures. Images were captured using a confocal laser scanning microscope. The lncRNA TCONS_00323213 FISH probe was synthesized by RiboBio, a biotechnology company(RiboBio, Guangzhou, China).
3
Subcellular Localization of lncRNA by FISH
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FISH assays were carried out with a lncRNA FISH Kit (RiboBio, Guangzhou, China). In brief, cells were fixed and permeabilized in PBS containing 0.5% Triton X-100. FISH probes were designed by RiboBio (Guangzhou, China). Hybridization was carried out overnight in a humidified chamber at 37 °C in the dark. All images were obtained with an Olympus FV1000 confocal microscope (Tokyo, Japan). 4′,6-diamidino-2-phenylindole and Cy3 channels were used to detect the signals. 18 S and U6 were used as the cytoplasmic and nuclear markers, respectively. Immunofluorescence staining was performed according to the manufacturer’s instructions, and an anti-PFKFB3 antibody (1:150, Abcam, ab181861) was used in this study.
4
Immunofluorescence Assay and RNA FISH for ARHGAP5-AS1
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The immunofluorescence assay was performed as previously reported.20 After permeabilisation and blockage, HepG2 cells were incubated with primary antibodies overnight. Cells were then stained with secondary antibodies (Table S1 ), washed with PBS and incubated with 4,6‐diamidino‐2‐phenylindole (DAPI). RNA FISH was performed to examine the co‐localization of lncRNA ARHGAP5‐AS1 and CSDE1 protein using the lncRNA FISH Kit (RiboBio) and immunofluorescence staining of CSDE1 antibody. In short, cells were permeabilised with 0.5% Triton X‐100 and hybridised with the FISH probes overnight at 37 °C in dark. LncRNA ARHGAP5‐AS1 signals were detected using Cy3 channels. CSDE1 was stained with its antibody and CoraLite488‐conjugated Goat Anti‐Rabbit IgG(H + L) (Table S1 ). A Zeiss LSM800 confocal microscope (Zeiss, Germany) was used to visualise images.
5
Comprehensive lncRNA FISH Assay Protocol
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FISH assays were carried out with a lncRNA FISH Kit (RiboBio, Guangzhou, China). In brief, cells and FISH probes were prepared. Then hybridization was conducted according to the protocol. All images were obtained with an Olympus FV1000 confocal microscope (Tokyo, Japan). Besides, immunofluorescence staining was proceed following to the manufacturer’s instructions, and an anti-STK33 antibody (1:150, Abcam, ab206296) was used in this study.
6
lncRNA Localization by FISH
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FISH was performed with the lncRNA FISH Kit (Guangzhou RiboBio, Guangzhou, China) according to the manufacturer’s instructions. Briefly, cells were fixed with 4% formaldehyde for 10 min at room temperature. After being washed three times with PBS, cells were permeabilized with 0.5% Triton X-100 for 10 min at 4°C. Then, cells were incubated with RNA probes in hybridization buffer overnight at 37°C. The RNA probes were directly conjugated with a fluorophore. The cells were then washed three times with saline sodium citrate buffer and stained with DAPI. Images were captured by confocal microscopy (LSM700META; Zeiss, Oberkochen, Germany).
7
Localization of lncRNA via FISH
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FISH assays were carried out with the lncRNA FISH Kit (RiboBio). In brief, the cells were fixed and permeabilized in PBS containing 0.5% Triton X-100. FISH probes were designed by RiboBio. Hybridization was carried out overnight in a humidified chamber at 37 °C in the dark. All images were obtained with a CarlZeiss LSM710 confocal microscope (Germany). 4’,6-Diamidino-2-phenylindole and Cy3 channels were used to detect the signals. U6 and 18S were used as the nuclear and cytoplasmic markers, respectively. Immunofluorescence staining was performed according to the manufacturer’s instructions. All antibody information is provided in Additional file 2 : Table S9.
8
lncRNA Visualization by Fluorescent In Situ Hybridization
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RNA fluorescence in situ hybridization (FISH) was performed using the lncRNA FISH Kit (Guangzhou RiboBio, China) according to the manufacturer's instructions. Briefly, cells were fixed with 4% formaldehyde for 10 min at room temperature. After washing, cells were permeabilized with 0.5% Triton X‐100 for 30 min at 37°C. Then, cells were incubated with RNA probes in hybridization buffer overnight at 37°C. The RNA probes were directly conjugated with a fluorophore. Then, the cells were washed three times using saline sodium citrate buffer, stained with 4,′6‐diamidino‐2‐phenylindole (DAPI) for 10 min at room temperature, and examined using a fluorescence microscope.
9
lncRNA Localization by FISH
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RNA fluorescence in situ hybridization (FISH) experiments were performed using an lncRNA FISH Kit (Guangzhou RiboBio, Guangzhou, China) according to the manufacturer's instructions. In brief, the cells were incubated with the RNA probes in hybridization buffer overnight at 37°C. The cells were washed three times with saline-sodium citrate buffer, stained with 4′,6-diamidino-2-phenylindole (DAPI) for 10 min at room temperature, and examined using a fluorescence microscope.
10
Co-localization of LINC00842 and PGC-1α in PDAC
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Co-localization of LINC00842 and PGC-1α protein was analyzed by RNA FISH performed with a lncRNA FISH Kit (RiboBio) and immunofluorescence staining of PGC-1α antibody. Briefly, PDAC cells were fixed and permeabilized in PBS containing 0.5% of Triton X-100. Hybridization with the FISH probes designed by RiboBio was carried out overnight in a humidified chamber at 37 °C in dark. 4′,6-diamidino-2-phenylindole and Cy3 channels were used to detect the signals. Immunofluorescence staining of PGC-1α was performed using PGC-1α antibody (1:200, NBP1-04676, Novus Biologicals, verification of antibody specificity was shown in Supplementary Fig. 15a ) and Alexa Fluor® 488 conjugated secondary antibody (1:500, A-21206, Invitrogen). Cell nuclei were counterstained with DAPI. Images were obtained with Olympus FV1000 confocal microscope.
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