Nuclear fast red

Manufactured by Vector Laboratories

Sourced in United States, Canada, United Kingdom, Germany

Nuclear Fast Red is a nuclear stain used in histology and cytology for the visualization of nuclei. It stains nuclei a bright red color, providing a clear contrast against the background. The dye binds to the phosphate groups of DNA, allowing for the identification and localization of cellular nuclei.

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Lab products found in correlation

Nbt bcip, by Roche (12 mentions) Senescence β galactosidase staining kit, by Cell Signaling Technology (6 mentions) Vectastain abc ap kit, by Vector Laboratories (5 mentions) Vectamount, by Vector Laboratories (5 mentions) Mgcl2, by Merck Group (4 mentions) Sheep anti dig ap, by Roche (4 mentions) Proteinase k, by Qiagen (4 mentions) Hybridization buffer, by Qiagen (3 mentions) Histoclear, by Avantor (3 mentions) Dpx mountant, by Merck Group (3 mentions) Vector blue ap substrate kit 3, by Vector Laboratories (3 mentions) 5 bromo 4 chloro 3 indolyl phosphate, by Roche (3 mentions) Of the substrate lactate, by Merck Group (3 mentions) Nitrotetrazolium blue chloride, by Merck Group (3 mentions) Phenazine methosulfate, by Merck Group (3 mentions) Proteinase k, by Merck Group (3 mentions) Nbt bcip, by Vector Laboratories (3 mentions) Superfrost plus slide, by Thermo Fisher Scientific (3 mentions) Alkaline phosphatase conjugated anti dig antibody, by Roche (3 mentions) Mircury lna microrna detection probe, by Qiagen (3 mentions)

Close spelling variants of this product

Fast Red (10 citations) Nuclear fast red counterstain (8 citations) Nuclear Fast Red solution (5 citations) Vector Nuclear Fast Red (4 citations) Nuclear Red (3 citations) Fast Red nuclear stain (2 citations) Nuclear fast red staining (2 citations)

154 protocols using nuclear fast red

Colonic Tissue Immunohistochemistry Protocol

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Colonic tissue was isolated and washed in phosphate buffer saline (PBS). Immunohistochemistry was performed as previously described [9 (link)]). Briefly, 4 mM paraffin sections were incubated with the corresponding primary antibody: ClC-2 1:400 (ACL-002, Alomone), NHE-3 1:1000 (C-20, Santa Cruz), and NKCC1 1:4000 (kindly provided by R. James Turner, [10 (link)]). To detect bound antibodies, we used the LSAB Universal Kit (Dako). The positive reaction was developed with diaminobenzidine or Vector SG (Vector Labs) and sections were counterstained with hematoxylin or nuclear fast red (Vector Labs).
For double immunohistochemistry, samples were first incubated with anti-ClC2 antibody, and peroxidase activity was visualized using Vector SG (Vector Labs). At the end of the first labeling reaction, sections were washed in 50 mM Tris HCl pH 7.8 and treated as described above for NHE3 immunolabeling. The counterstaining was performed with nuclear fast red (Vector Labs).

Millar-Büchner P., Philp A.R., Gutierrez N., Villanueva S., Kerr B, & Flores C.A. (2016). Severe changes in colon epithelium in the Mecp2-null mouse model of Rett syndrome. Molecular and Cellular Pediatrics, 3, 37.

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Histological Analysis of Testis and Lung

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P14 mice were sacrificed, and the testes and lungs were dissected. After three washes with phosphate-buffered saline (PBS), the tissues were fixed in 4% paraformaldehyde (PFA)/0.02% Nonidet P-40 (NP-40) for 2 h at room temperature and permeabilized with 0.02% NP-40 in PBS for 1 h. Samples were incubated overnight at 37 °C in X-gal staining solution composed of 5 mM K₃Fe(CN)₆, 5 mM K₄Fe(CN)₆, 2 mM MgCl₂, 0.01% sodium deoxycholate, 0.02% NP-40, and 1 mg/mL X-gal in PBS. They were then washed three times with PBS for 5 min each wash and post-fixed with 4% PFA for 24 h before being embedded in paraffin. Sections (10 μm thick) were deparaffinized and rehydrated through a graded series of ethanol concentrations followed by counterstaining with Nuclear Fast Red (Vector Laboratories, Burlingame, CA, USA).

Kim D.Y., Sub Y.J., Kim H.Y., Cho K.J., Choi W.I., Choi Y.J., Lee M.G., Hildebrandt F, & Gee H.Y. (2023). LRRC6 regulates biogenesis of motile cilia by aiding FOXJ1 translocation into the nucleus. Cell Communication and Signaling : CCS, 21, 142.

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Functional Enhancer Assays of Chimpanzee GDF6

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The chimpanzee versions of sequences missing in the human GDF6 locus were PCR amplified and cloned into hsp-lacZ and hsp-CreER-T2 minimal promoter expression vectors and injected into the pronuclei of fertilized FVB embryos (Xenogen and Stanford Transgenic Facility). The hCONDEL.306-hsp-CreER-T2 transgenic mice were then bred to floxed-ROSA26 reporter mice, and injected with tamoxifen at different developmental time points. For both types of constructs, functional enhancer assays were then carried out by staining for lacZ expression activity at multiple developmental stages. For more detailed analysis, lacZ stained hindlimbs were embedded in gelatin, sectioned in transverse orientation using a Leica CM3050 S, and counterstained with Nuclear Fast Red (Vector Laboratories, CA). See Extended Experimental Procedures for further details.

Indjeian V.B., Kingman G.A., Jones F.C., Guenther C.A., Grimwood J., Schmutz J., Myers R.M, & Kingsley D.M. (2016). Evolving New Skeletal Traits by cis-Regulatory Changes in Bone Morphogenetic Proteins. Cell, 164(0), 45-56.

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miRNA Detection in FFPE Tissues

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miRCURY locked nucleic acid (LNA)™ microRNA detection probes, namely, rno-miR-21, rno-miR-31, rno-miR-223, hsa-miR-31, hsa-miR-223, negative controls (rno-miR-31) with mismatches at two position, were purchased from Exiqon (Vedbaek, Denmark). The oligonucleotides are double DIG-labeled at the 5′- and 3′-ends. ISH was performed on 6 μm FFPE sections as described by Nielsen et al. [91 (link)]. Following deparaffinization, rehydration in graded alcohol and proteinase K treatment, tissue sections were hybridized with miR-31 probe (20 nM), miR-223 or miR-21 probe (50 nM) in hybridization buffer (Exiqon) at 50°C - 57°C for 14 h in a hybridizer (Dako, Glostrup, Denmark). Following stringent washes in SSC buffers, the sections were blocked against unspecific binding of the detecting antibody, using DIG wash and blocking reagent. miRNA was localized by incubation with 4-nitro-blue tetrazolium (NBT) and 5-brom-4-chloro-3′-indolylphosphate (BCIP) (Roche, Mannheim, Germany). Nuclear fast red (Vector Lab., Burlingname, CA) was used as a counterstain.

Fong L.Y., Taccioli C., Jing R., Smalley K.J., Alder H., Jiang Y., Fadda P., Farber J.L, & Croce C.M. (2016). MicroRNA dysregulation and esophageal cancer development depend on the extent of zinc dietary deficiency. Oncotarget, 7(10), 10723-10738.

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Thrombus Characterization in Inferior Vena Cava

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The thrombosed IVC was excised from the site of stenosis to the iliac bifurcation at 1, 7 and 14 days post-induction. Samples were fixed in 10% formol saline for 18 h and processed in a Leica TP1020 tissue processor (Leica, Germany). 5 μm paraffin sections were cut at 500 μm intervals throughout the length of the thrombus. Haematoxylin and eosin (H&E) staining was used to visualise the thrombus within the IVC as previously described [12] (link). Perls's staining was used to identify intracellular iron deposits in thrombus sections by incubation with a 2%(w/v) potassium ferrocyanide in 2%(v/v) hydrochloric acid solution for 20 min, followed by a distilled water wash and counterstaining for 5 min with Nuclear Fast Red (Vector Laboratories, UK). All sections were dehydrated and mounted in DPX medium (Sigma, USA).

Grover S.P., Saha P., Jenkins J., Mukkavilli A., Lyons O.T., Patel A.S., Sunassee K., Modarai B, & Smith A. (2015). Quantification of experimental venous thrombus resolution by longitudinal nanogold-enhanced micro-computed tomography. Thrombosis Research, 136(6), 1285-1290.

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In Situ β-Galactosidase Assay on Cryosections

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Freshly obtained tissues were embedded in optimal cutting temperature compound (Tissue Tek). Cryosections were done at 5 μm on a cryostat set at −25°C. For in situ β-galactosidase activity assays, sections were fixed for 10 min at 4°C in 0.5% glutaraldehyde/PBS. Samples were then washed with PBS three times for 5 min each and stained for 6–24 hr at 37°C with X-gal working solution with 1 mg/mL X-gal in PBS, 5 mM K₃Fe(CN)₆, and 5 mM K₄Fe(CN)₆, and counterstained with nuclear fast red (Vector Labs) or eosin (Sigma) (Folgueras et al., 2013 (link)).

Lin C.L., Xu R., Yi J.K., Li F., Chen J., Jones E.C., Slutsky J.B., Huang L., Rigas B., Cao J., Zhong X., Snider A.J., Obeid L.M., Hannun Y.A, & Mao C. (2017). Alkaline Ceramidase 1 Protects Mice from Premature Hair Loss by Maintaining the Homeostasis of Hair Follicle Stem Cells. Stem Cell Reports, 9(5), 1488-1500.

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X-gal Staining and Imaging Protocol

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Sections were thawed at room temperature and rehydrated in PBS supplemented with 2 mm MgCl₂ for 5 min before being incubated overnight at 37°C in X‐gal staining solution (PBS (pH 7.4) containing 2 mm MgCl₂, 0.01% (w/v) sodium deoxycholate, 0.02% (v/v) Igepal CA630, 5 mm potassium ferricyanide, 5 mm potassium ferrocyanide and 1 mg ml⁻¹ 5‐bromo‐4‐chloro‐3‐indolyl β‐d‐galactopyranoside). On the following day, slides were washed in PBS, counterstained in Nuclear Fast Red (Vector Laboratories) for 5 min, washed twice in distilled water and dehydrated through 70% and 95% ethanol before being incubated in Histoclear (VWR) for 3 min, air‐dried and mounted in DPX mountant (Sigma). Slides were digitized at 40× magnification using an Aperio digital scanner (Leica) and TIF images were extracted using ImageScope software (Leica). The resulting images were rendered in ImageJ.

McMahon M., Ding S., Acosta‐Jimenez L.P., Frangova T.G., Henderson C.J, & Wolf C.R. (2017). Measuring in vivo responses to endogenous and exogenous oxidative stress using a novel haem oxygenase 1 reporter mouse. The Journal of Physiology, 596(1), 105-127.

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In situ Hybridization of miRNAs in CRC Tumors

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For in situ hybridization of miRNAs, 13 CRC tumours were used. Sections (5 µm) were cut in RNAse free water, deparaffinized, incubated with Proteinase-K (10 µg/ml) for 15 min at 37 °C, dehydrated and air-dried. Double-DIG-labelled miRNA probes (Exiqon, Denmark): miR-210 (40 nM) and scrambled (60 nM). Incubation was at 56 °C for 2 h with 25 µL of probe followed by washing with saline–sodium citrate. Slides were incubated with blocking buffer for 15 min, then with anti-DIG-Alkaline phophatase reagent (1:600, Exiqon, Denmark) at RT. Slides were washed with PBS–Tween and incubated with anti-AP substrate (NCT-BCIP, Roche, UK) for 2 h at RT, washed in water before counterstaining with Nuclear Fast Red (Vector, UK). Slides were rinsed in running tap water for 10 min, dehydrated, mounted with Eukkitt mounting medium (Sigma, UK) and then scanned using a Hamamatsu slide scanner.

Nijhuis A., Thompson H., Adam J., Parker A., Gammon L., Lewis A., Bundy J.G., Soga T., Jalaly A., Propper D., Jeffery R., Suraweera N., McDonald S., Thaha M.A., Feakins R., Lowe R., Bishop C.L, & Silver A. (2017). Remodelling of microRNAs in colorectal cancer by hypoxia alters metabolism profiles and 5-fluorouracil resistance. Human Molecular Genetics, 26(8), 1552-1564.

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Whole-mount β-galactosidase staining of embryos

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Whole-mount staining for β-galactosidase activity was performed as previously described (Capellini et al., 2017 (link)). Gestational day (E) 14.5 R4 enhancer lacZ positive embryos were hemisected and fixed in 4% paraformaldehyde (PFA) (Sigma, 158127) at 4°C, according to gestational-day guidelines. Fixed embryos were washed three times in wash buffer and stained for 16–24 hours in the dark with 1 mg/ml X-gal (Sigma, B4252) in staining buffer at room temperature. Adult R4 enhancer lacZ positive mice were skinned and eviscerated, and the limbs were removed. Each limb then had its most superficial muscles removed and its joint capsules gently pierced to permit solutions from entering into the joint cavity. Fixation and staining times were adjusted accordingly for these post-natal specimens. After staining, embryos and post-natal limbs were briefly rinsed in wash buffer and post-fixed in 4% PFA at 4°C for 5 hours. For sectioning, X-gal stained embryos were placed first in sucrose and then embedded in gelatin/sucrose solution and cryo-sectioned at 25 μm. Sections were counterstained with Nuclear Fast Red (Vector labs, #H-3403).

Richard D., Liu Z., Cao J., Kiapour A.M., Willen J., Yarlagadda S., Jagoda E., Kolachalama V.B., Sieker J.T., Chang G.H., Muthuirulan P., Young M., Masson A., Konrad J., Hosseinzadeh S., Maridas D.E., Rosen V., Krawetz R., Roach N, & Capellini T.D. (2020). Evolutionary Selection and Constraint on Human Knee Chondrocyte Regulation Impacts Osteoarthritis Risk. Cell, 181(2), 362-381.e28.

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Histochemical Analysis of β-Galactosidase Activity

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Sections were thawed at room temperature and rehydrated in PBS supplemented with 2 mM MgCl₂ for 5 minutes before being incubated overnight at 37°C in X-gal staining solution (PBS (pH 7.4) containing 2 mM MgCl₂, 0.01% (w/v) sodium deoxycholate, 0.02% (v/v) Igepal CA630, 5 mM potassium ferricyanide, 5 mM potassium ferrocyanide and 1 mg/ml 5-bromo-4-chloro-3-indolyl β-D-galactopyranosidise). On the following day, slides were washed in PBS, counterstained in Nuclear Fast Red (Vector Laboratories) for 5 minutes, washed twice in distilled water and dehydrated through 70% and 95% ethanol before being incubated in Histoclear (VWR) for 3 minutes, air-dried and mounted in DPX mountant (Sigma). Sections were evaluated by an expert, independent clinical pathologist (Dr Shaun Walsh, Department of Pathology, Ninewells Hospital, Dundee). Dr Walsh was blinded in the case of sections from olaparib-treated mice (Fig 5). Brain sections were additionally examined by a neuroscientist (Dr John Sharkey, University of Dundee). Images were captured using a Zeiss 12 megapixel digital camera attached to a Zeiss Axio Scope.A1 microscope, and controlled by means of the AxioVision v4.5 software (Carl Zeiss). Images were rendered in ImageJ.

McMahon M., Frangova T.G., Henderson C.J, & Wolf C.R. (2016). Olaparib, alone or in combination with ionizing radiation, exacerbates DNA damage in normal tissues, as revealed by a new p21 reporter mouse. Molecular cancer research : MCR, 14(12), 1195-1203.

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