Cell cycle profiles were obtained by adding 10 μM of 5-bromo-2′-deoxyuridine (BrdU, Sigma-Aldrich) to HeLa cells 1 hr prior to harvesting. Single-cell suspensions were washed with PBS, fixed in ice-cold 70% ethanol and kept overnight at 4°C. For BrdU staining, cells were incubated in 2N HCl, 0.5% Triton X-100 for 30 min at room temperature followed by a 2 min incubation in 0.1 M sodium tetraborate buffer, pH 8.0. Cells were then washed with PBS, 1% BSA and incubated with FITC-conjugated anti-BrdU antibody (BioLegend) in PBS, 1% BSA, 0.5% Tween-20 for 1 hr at room temperature. They were then washed again in PBS and stained with 0.02 mg/mL propidium iodide (PI) in PBS, 0.1% Triton X-100, 0.2 mg/mL RNase A for 30 min at room temperature. Samples were acquired on a FACSCalibur flow cytometer (BD Biosciences) and analyzed with FlowJo software (Tree Star).
Fitc conjugated anti brdu antibody
Manufactured by BioLegend
Sourced in United States
FITC-conjugated anti-BrdU antibody is a laboratory reagent used for the detection of bromodeoxyuridine (BrdU) incorporation into cellular DNA. It is a fluorescently labeled antibody that binds specifically to BrdU, allowing for the visualization and quantification of cellular proliferation.
Automatically generated - may contain errors
Lab products found in correlation
5 bromo 2 deoxyuridine brdu, by Merck Group (2 mentions)
Bromodeoxyuridine (brdu), by Merck Group (2 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Canto 2, by BD (1 mentions)
Propidium iodide, by Serva Electrophoresis (1 mentions)
Idasanutlin, by Selleck Chemicals (1 mentions)
Facsverse flow cytometer, by BD (1 mentions)
Rnase a, by Thermo Fisher Scientific (1 mentions)
Modfit lt software, by Verity Software House (1 mentions)
Bromodeoxyuridine (brdu), by BioLegend (1 mentions)
Facscalibur, by BD (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Nacalai Tesque (1 mentions)
5 bromo 2 deoxyuridine brdu, by Nacalai Tesque (1 mentions)
Attune nxt flow cytometer, by Thermo Fisher Scientific (1 mentions)
Carboxyfluorescein succinimidyl ester (cfse), by BioLegend (1 mentions)
8 protocols using fitc conjugated anti brdu antibody
1
Cell Cycle Analysis by BrdU and PI
2
BrdU Incorporation Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were fixed and treated with DNase, followed by incubation with FITC‐conjugated anti‐BrdU antibody according to instructions (Biolegend, Cat#364103). The flow cytometry assay was performed using a Canto II flow cytometer (BD Bioscience). The data were analyzed using FlowJo v10.0 software (Tree Star).
3
Cell Cycle Analysis with BrdU Labeling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The cells were treated with DMSO or idasanutlin (RG7388, Selleck Chemicals, Houston, TX, USA) for 24 h with pulse-labeling using 10 µM bromodeoxyuridine (BrdU, Sigma Aldrich) for the last hour of the treatment. After that, the cells were harvested by trypsinization, fixed with 96% ethanol and stored at −20 °C. The cells were centrifuged and suspended in 2 M HCl/0.5% Triton X-100, added dropwise. Following 30 min incubation at room temperature and centrifugation, the cells were resuspended in 0.1 M Na2B4O7, pH 8.5, centrifuged again and suspended in 0.5% Tween 20/1% BSA/PBS. FITC-conjugated anti-BrdU antibody (BioLegend, cat. 364104, San Diego, CA, USA) was then added (5 µL per 106 cells) and the cells were incubated at 4 °C overnight with gentle shaking (200 rpm). The cells were centrifuged, suspended in PBS containing 5 µg/mL of propidium iodide (PI, SERVA Electrophoresis GmbH, Heidelberg, Germany) and 10 µg/mL RNase A (Thermo Scientific, Waltham, MA, USA), incubated for 20 min at room temperature, and analyzed with BD FACSVerse flow cytometer (Becton Dickinson, Franklin Lakes, NJ, USA). Cell cycle distribution was analyzed using ModFit LT Software (Verity Software House, Topsham, ME, USA).
4
BrdU Labeling of Proliferating Cells
5
CXCR5 Silencing Impacts Cell Proliferation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were plated in 96-well plates at a density of 6×103 cells/well. The cells were transfected 100 nmol/l human CXCR5 siRNA or control siRNA using Lipofectamine 2000 for 6 h, and cell proliferation was analysed using a CCK-8 assay according to the manufacturer's instructions. The absorbance at 490 nm was then obtained using an enzyme-linked immunosorbent assay plate reader. To stain proliferating cells, LNCaP cells were stained with a FITC-conjugated anti-BrdU antibody (Biolegend) as well as DAPI. At least 150 cells were counted per condition, and the percentage of BrdU-positive cells is shown.
6
Cell Cycle Analysis by Flow Cytometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Synchronised HeLa cells were washed with PBS buffer and fixed in ice-cold ethanol overnight. DNA was stained with a propidium iodide (PI) solution containing 0.1% Triton X-100 in PBS, 0.2 mg/ml RNase A, 0.02 mg/ml PI for 30 min at room temperature. Alternatively, BrdU-treated cells were fixed by adding ice-cold 70% ethanol overnight. Cells were then incubated with 2 N HCl, 0.5% Triton X-100 for 30 min at room temperature and washed with 0.1 M sodium tetraborate buffer, pH 8 for 2 min. Cells were washed and incubated with a FITC conjugated anti-BrdU antibody (BioLegend) for 1 hr at room temperature. PI staining followed as described above. Cell cycle profiles were analysed by flow cytometry with FACSCalibur (BD Biosciences) and processed using FlowJo software (Tree Star).
7
Cell Cycle Analysis by Flow Cytometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell cycle was determined by flow cytometry of cells labelled with anti-BrdU-FITC and DAPI. Asynchronously growing cells were pulse-labelled with 10 µM 5- bromo-2′-deoxyuridine (BrdU) (Nacalai Tesque) for 3 h prior to harvesting. Cells were trypsinized and fixed in ice-cold 70% ethanol for 30 min on ice. Cells were suspended in 2 M HCl containing 0.5% (w/v) Triton X-100 for 30 min at room temperature. After supernatant was discarded, cells were resuspended in 0.1 M Na2B4H7 (pH 8.5) for 2 min at room temperature. Cells were then washed in buffer (1% bovine serum albumin (BSA) and 0.05% Tween 20 containing PBS) and treated with FITC-conjugated anti-BrdU antibody (BioLegend, San Diego, CA, USA) overnight at 4 °C. Cells were washed with 1% BSA containing PBS, centrifuged, and resuspended in 10 µg/mL 4′,6-diamidino-2-phenylindole (DAPI) (Nacalai Tesque), and allowed to incubate for 30 min on ice. Cell cycle analysis was performed on the Attune NxT flow cytometer (Thermo Fisher Scientific).
8
Cell Proliferation Analysis by CFSE and BrdU
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For analysis of cell proliferation, 10x106 cells were stained with 2.5 μM carboxyfluorescein succinimidyl ester (CFSE, Biolegend) in 2 mL staining buffer (0.1% BSA in PBS at 37°C) for 10 min at room temperature. The reaction was stopped by adding cells to 40 mL ice-cold RPMI-1640 culture medium. Cells were washed twice and cultured for 3 days before FACS analysis. Cell cycle profiles were obtained by adding 10 μM of 5-bromo-2′-deoxyuridine (BrdU, Sigma-Aldrich) to 0.5-1x106 CH12 cells 1 hr before harvesting. Cells were washed in PBS, fixed in ice-cold 70% ethanol and kept overnight at 4°C. For BrdU staining, cells were incubated in 2N HCl, 0.5% Triton X-100 for 30 min at room temperature followed by a 2 min incubation in 0.1 M sodium tetraborate buffer, pH 8.0. Cells were washed in PBS, 1% BSA and incubated with 2 μL FITC-conjugated anti-BrdU antibody (BioLegend) diluted in PBS, 1% BSA and 0.5% Tween-20 for 1 hr at room temperature. Cells were washed once and stained with 0.02 mg/mL propidium iodide (PI) in PBS, 0.1% Triton X-100, 0.2 mg/mL RNase A for 30 min at room temperature before FACS analysis.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!