Bromodeoxyuridine (brdu)

Manufactured by Bio-Rad

Sourced in United Kingdom, United States

BrdU is a nucleoside analog that can be incorporated into the DNA of dividing cells, serving as a marker for cell proliferation. It can be detected using specific antibodies.

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Lab products found in correlation

Bromodeoxyuridine (brdu), by Merck Group (7 mentions) Cleaved caspase 3, by Cell Signaling Technology (6 mentions) Neuronal nuclei (neun), by Merck Group (5 mentions) Rat anti brdu antibody, by Bio-Rad (2 mentions) β catenin, by Cell Signaling Technology (2 mentions) 5 ethynyl 2 deoxyuridine (edu), by Thermo Fisher Scientific (2 mentions) Glial fibrillary acidic protein (gfap), by Merck Group (2 mentions) Nanoinjector, by World Precision Instruments (1 mentions) Infinite horizon impactor, by Precision Systems and Instrumentation (1 mentions) Nod scid mice, by Jackson ImmunoResearch (1 mentions) γh2ax, by Cell Signaling Technology (1 mentions) P27kip1, by Agilent Technologies (1 mentions) Phospho histone h3, by Cell Signaling Technology (1 mentions) Proliferating cell nuclear antigen (pcna), by Cell Signaling Technology (1 mentions) Aperio scanscope xt, by Leica (1 mentions) Click it chemistry, by Thermo Fisher Scientific (1 mentions) 5 ethynyl 2 deoxyuridine edu, by Thermo Fisher Scientific (1 mentions) 5 bromo 2 deoxyuridine brdu, by Merck Group (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions) Dnase 1, by Merck Group (1 mentions)

29 protocols using bromodeoxyuridine (brdu)

Proliferation of Human OPCs

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Human CD140a⁺ OPCs were infected with lentivirus as described above and cultured for 48 h in SFM supplemented with 20 ng/ml PDGF-AA and 5 ng/ml NT-3. BrdU (30 μM, Sigma) was then added, and cells were fixed 24 h later. Cells were immunostained for BrdU (1:1000; AbD Serotec) and the percentage of BrdU⁺ DAPI-stained nuclei was obtained from 5–6 fields for each fetal sample.

Wang J., Saraswat D., Sinha A.K., Polanco J., Dietz K., O’Bara M.A., Pol S.U., Shayya H.J, & Sim F.J. (2018). Paired Related Homeobox Protein 1 Regulates Quiescence in Human Oligodendrocyte Progenitors. Cell reports, 25(12), 3435-3450.e6.

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BrdU Labeling of Proliferating Cells

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The mice were given a dose of BrdU (Sigma, Cat: B5002) intraperitoneal at 100mg/kg 24 hours before sacrifice. After rinsing in 0.01M PBS, the specimens were treated with 2 N HCl for 30 min at 37°C for partial denaturalization of double-stra nded DNA. To reveal BrdU, the specimens were incubated with a rat anti-BrdU (1:200, BIO-RAD, Cat: OBT0030) overnight, then the sections were incubated with Alexa 488-conjugated secondary antibody.

Wang F., Yang Y.J., Yang N., Chen X.J., Huang N.X., Zhang J., Wu Y., Liu Z., Gao X., Li T., Pan G.Q., Liu S.B., Li H.L., Fancy S.P., Xiao L., Chan J.R, & Mei F. (2018). Enhancing oligodendrocyte myelination rescues synaptic loss and improves functional recovery after chronic hypoxia. Neuron, 99(4), 689-701.e5.

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Contusion SCI Treated with hCNS-SCns Transplant

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Contusion SCIs followed by early chronic hCNS-SCns transplantation into the intact parenchyma were performed as described previously (Cummings et al. 2005 (link); Salazar et al. 2010 ). Briefly, adult female NOD-scid mice (Jackson Laboratory, Sacramento, CA) were anesthetized with isoflurane (VetEquip Inc., Pleasanton, CA), received a T9 laminectomy using a surgical microscope, and a bilateral 50-kDa contusion injury using the Infinite Horizon Impactor (Precision Systems and Instrumentation, Lexington, KY). Thirty days post-SCI, the mice were re-anesthetized and 1.25 μl of freshly triturated hCNS-SCns suspension were injected at four bilateral sites (for a total of 5 μl) 0.75mm from midline. Two injection sites were at the posterior aspect of T8 (rostral to the site of injury), and two at the anterior aspect of T10 (caudal to the site of injury). Injections were conducted using a Nanoinjector with a micro controller (World Precision Instruments, Waltham, MA) at speed of 417 nl/minute, followed by a 2 min delay before withdrawal of the needle, using pulled silicon-treated glass injection tips with a 70μm ID and 110μm OD (Sutter Instruments, Novato, CA). For assessing hCNS-SCns proliferation at 2 DPT or 2 WPT, the mice were injected i.p. with 50 mg/kg of BrdU (AbD Serotec, Raleigh, NC) every 12 hr starting from the time of transplantation until 2 DPT or 2 WPT.

Piltti K.M., Avakian S.N., Funes G.M., Hu A., Uchida N., Anderson A.J, & Cummings B.J. (2015). Transplantation dose alters the dynamics of human neural stem cell engraftment, proliferation and migration after spinal cord injury. Stem cell research, 15(2), 341-353.

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Immunohistochemical Analysis of Cell Markers

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IHC was carried out in the Bond stainer (Leica). In brief, slides were dewaxed in Bond Dewax solution) and hydrated in bond wash solution. Antigen retrieval for antibodies was performed for 30 min at 100°C in bond-epitope retrieval solution 1 (pH 6.0). Slides were incubated with primary antibody for 1 hr. Primary antibodies used were Dicer 13D6 (Abcam), cleaved caspase-3 (Cell Signaling Technology), γH2AX (Cell Signaling Technology), p27-Kip1 (Dako), PCNA (Cell Signaling Technology), BrdU (AbD Serotec), Ki-67 (Leica), phospho-histone H3 (Cell Signaling Technology), and NeuN (Millipore). Nuclei were counterstained with hematoxylin or DAPI. Antibody detection was performed using the Bond Polymer Refine Detection System (DS9800). Stained slides were dehydrated and coverslipped. Stained slides were digitally imaged at 20× magnification using the Aperio ScanScope XT (Aperio Technologies), and digital images were stored in the Aperio eSlide Manager Database at Translation Pathology Laboratory (TPL).

Swahari V., Nakamura A., Baran-Gale J., Garcia I., Crowther A.J., Sons R., Gershon T.R., Hammond S., Sethupathy P, & Deshmukh M. (2015). Essential Function of Dicer in Resolving DNA Damage in the Rapidly Dividing Cells of the Developing and Malignant Cerebellum. Cell reports, 14(2), 216-224.

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Visualizing DNA Replication in C2C12 Cells

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For the visualization of replicating DNA, C2C12 mouse myoblasts were pulse labelled for 30 min with 100 μM 5-bromo-2′-deoxyuridine, BrdU (Sigma-Aldrich, Steinheim, Germany, Cat #: 59-14-3) and/or 10 μM 5′-ethynyl-2′-deoxyuridine, EdU (Invitrogen, Carlsbad, USA, Cat #: C10339). For a pulse-chase-pulse experimental setup a 200 μM thymidine chase (Sigma-Aldrich, Steinheim, Germany, Cat #: T1895) was performed in between both pulses. Incorporated BrdU was detected with a rat anti-BrdU antibody (1/200, AbD Serotec, Oxford, UK, Cat #: OBT0030CX) combined with 10 μg/μl DNaseI (Sigma-Aldrich, Steinheim, Germany, Cat #: D5025) for 1 h at 37°C in 4% BSA/PBS. Cells were then washed with 0.5% BSA/1mM EDTA/PBS + 0.01% Tween to stop DNaseI digestion. EdU was detected using ClickIT chemistry (Invitrogen, Carlsbad, USA, Cat #: C10639) as described in (41 (link)) with Alexa Fluor 594 (1/300). DNA was counterstained with 1 μg/ml DAPI (Sigma-Aldrich, Steinheim, Germany, Cat #: D9542) for 10 min at RT and cells were mounted in Mowiol.

Heinz K.S., Casas-Delucchi C.S., Török T., Cmarko D., Rapp A., Raska I, & Cardoso M.C. (2018). Peripheral re-localization of constitutive heterochromatin advances its replication timing and impairs maintenance of silencing marks. Nucleic Acids Research, 46(12), 6112-6128.

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Immunohistochemical Analysis of Colon Tissues

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Immunohistochemical analysis was conducted on formalin‐fixed paraffin‐embedded tissues. Colon sections were stained with antibodies specific for β‐catenin (Cell Signaling Technology), BrdU (AbD Serotec), cleaved caspase‐3 (Cell Signaling Technology), CD31 (Dianova), c‐MYC (Santa Cruz Biotechnology), cyclin D1 (Santa Cruz Biotechnology), F4/80 (Life Technologies), iNOS (Abcam), and Ki‐67 (Dako Deutschland GmbH). The antibody‐antigen complexes were detected using biotinylated donkey anti‐rat and donkey anti‐rabbit secondary antibodies (Dianova) and the Dako REAL Detection System (Dako). Immunohistochemical detection of HIF‐1α (Cayman Chemical) was performed as previously described.23 Nuclei were counterstained with hematoxylin. Negative controls were performed by omitting the primary antibody. The average number of positively stained cells within at least six high power fields (HPF, 0.237 mm²) was determined by a blinded independent investigator.

Rohwer N., Kühl A.A., Ostermann A.I., Hartung N.M., Schebb N.H., Zopf D., McDonald F.M, & Weylandt K. (2020). Effects of chronic low‐dose aspirin treatment on tumor prevention in three mouse models of intestinal tumorigenesis. Cancer Medicine, 9(7), 2535-2550.

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Quantification of Neural Stem Cells

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Immunostaining was performed on a complete series of 40-μm sections, 240 μm apart, using primary antibodies against BrdU (1:500; AbD Serotec), Ki67 (1:1,000; eBioscience), and DCX (1:500; Santa Cruz; see also Table S4). Counting was performed blinded to the experimental groups at 400× magnification.

Leiter O., Seidemann S., Overall R.W., Ramasz B., Rund N., Schallenberg S., Grinenko T., Wielockx B., Kempermann G, & Walker T.L. (2019). Exercise-Induced Activated Platelets Increase Adult Hippocampal Precursor Proliferation and Promote Neuronal Differentiation. Stem Cell Reports, 12(4), 667-679.

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Immunohistochemical Analysis of Neuronal Markers

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Larvae were fixed in 4% paraformaldehyde and 0.025% glutaraldehyde in PBS at pH 7.4 for 1–2 h at 4° C, rinsed twice in Dulbecco’s calcium–magnesium-free PBS, cryoprotected in 30% sucrose and embedded in optimal cutting temperature medium (Tissue-Tek) for cryostat sections. 10 μm sections were taken following an anterior to posterior progression through the forebrain. One or 2 night incubation at 4°C with primary antibodies to TH (Imgenex), GABA (Millipore), Doublecortin (Abcam), Pax6 (Covance); BrdU (AbD Serotec); VGLUT 1,2 (Synaptic Systems), nNOS (Millipore), and c-Fos (Santa Cruz Biotechnology, Inc.) was followed by incubation with fluorescently tagged secondary antibodies for 1 hr 30 min at 20–22°C. Immunoreactivity of stained sections was examined on a SP5 Leica confocal system; z-stacks were acquired to confirm colocalization and to generate through-series projections. TH, the rate-limiting enzyme for DA synthesis, was used as a marker for DA and confirmed by DA immunostaining.

Dulcis D., Lippi G., Stark C.J., Do L.H., Berg D.K, & Spitzer N.C. (2017). Neurotransmitter Switching Regulated by miRNAs Controls Changes in Social Preference. Neuron, 95(6), 1319-1333.e5.

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Immunohistochemical Analysis of Lung Tissues

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Lung tissues sections were prepared as described previously (9 (link), 20 (link)). In addition, endogenous peroxidase activity was quenched with 3%(v/v) H₂0₂ in methanol for 20 min and then blocked in either 5% goat serum or serum-free block (DAKO) for 30 min at RT. Antibodies were applied overnight at 4C: CC10, SP-C and TTF1 (Santa Cruz); BrdU (Serotec); p19^Arf and Ki67 (Abcam); pT202/Y204-ERK1/2, p53 (Novocastra), p21 (Abcam) and p27^KIP1 (CST). The appropriate secondary was applied followed either by an anti-peroxidase or ABC amplification for 3–60 minutes at room temperature. Antigen-antibody complexes were detected using a DAB chromagen system (Sigma). PBS with 0.1%(v/v) Tween 20 was used for all washes.

Shai A., Dankort D., Juan J., Green S, & McMahon M. (2015). TP53 Silencing Bypasses Growth Arrest of BRAFV600E-Induced Lung Tumor Cells In a Two-Switch Model of Lung Tumorigenesis. Cancer research, 75(15), 3167-3180.

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Immunohistochemical Analysis of Transplanted Cells

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Immunohistochemistry was performed on coronal sections (16 μm) of mouse forebrain, as previously described (Sim et al., 2011 (link)). Human cells were identified with mouse anti-human nuclei antibodies (hNA; 1:400; Millipore), and co-labeled with markers for oligodendrocytes (CC1, 1:50; OLIG2, 1:750; Millipore), astrocytes (GFAP, SMI21; 1:1000; Covance) and OPCs (hNG2, 9.2.27; 1:200; Millipore). Overexpression was detected using anti-PRRX1 (1:300; OriGene) and anti-FLAG (1:200; Sigma) antibodies. Proliferating cells were detected using antibodies against Ki67 (1:25; BD Biosciences) or BrdU (1:2000; AbD Serotec). Alexa Fluor (488, 594, and 647) secondary antibodies (ThermoFisher Scientific) were used at 1:500 dilution. Sections were imaged with a 20 × objective using an inverted fluorescence microscope (Olympus IX51). The quantification of hNA⁺ and double-positive cells was performed by counting cells in midline and lateral regions of the corpus callosum; images from at least 9 fields containing >200 cells were collected per animal. To assess the migration of implanted cells, sections spanning from bregma +3.9 to −3.4 mm (Paxinos and Franklin, 2004 ) were sampled every 160 μm and immunostained for hNA. The rostrocaudal migration distance was calculated as the distance between the first and last sections containing hNA⁺ cells.

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