5-BrdU (Synonyms: BrdU, Bromodeoxyuridine, Broxuridine, NSC 38297) |
| رقم الكتالوجGC11940 |
مشتق صناعي للثايمدين
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 59-14-3
Sample solution is provided at 25 µL, 10mM.
;)
,-1-7$$$37316672.jpg');)
;)
;)
$$$36806353.jpg');)
;33(7)%EF%BC%9A546-561$$$37156877.jpg');)
;)
;)
;)
%201124-1138.$$$35908550.jpg');)
%20766-780.$$$37100057.jpg');)
%20418-432.$$$36893736.jpg');)
%EF%BC%9A-240-255.$$$35108512.jpg');)
533-545$$$36543525.jpg');)
%201-13.$$$36600053.jpg');)
;)
5-BrdU (5-bromo-2-deoxundine5-BrdU) is a synthetic thymidine bromide analog that is commonly used to measure DNA synthesis and label dividing cells [1]. 5-BrdU can selectively insert into cellular DNA in place of thymidine during the S phase and can be used to study signaling pathways and other physiological processes that induce cell proliferation [2]. 5-BrdU can be probed by in vitro cell culture or in vivo injection and then specifically detected using anti-BdU antibodies [3]. After 5-BrdU labeling, an additional DNA denaturation step is required after fixation and permeabilization of tissues or cells to allow anti-5-BrdU antibodies to bind to 5-BrdU incorporated into DNA [4]. 5-BrdU antibodies can be used in combination with other cell markers such as Ki67, doublecortin (DCX) and NeuN to identify proliferating cells and newly differentiated neurons [5].
References:
[1] Zhu H. Cell proliferation assay by flow cytometry (BrdU and PI staining)[J]. Bio-protocol, 2012: e198-e198.
[2] Ziv Y, Ron N, Butovsky O, et al. Immune cells contribute to the maintenance of neurogenesis and spatial learning abilities in adulthood[J]. Nature neuroscience, 2006, 9(2): 268-275.
[3] Matiašová A, Ševc J, Mikeš J, et al. Flow cytometric determination of 5-bromo-2ʹ-deoxyuridine pharmacokinetics in blood serum after intraperitoneal administration to rats and mice[J]. Histochemistry and cell biology, 2014, 142: 703-712.
[4] Wolter S, Dittmar F, Seifert R. cCMP and cUMP in apoptosis: concepts and methods[J]. Non-canonical Cyclic Nucleotides, 2017: 25-47.
[5] Diaz F. Characterization of the Neurogenic Response to Apoptosis of Cortical Glutamatergic Neurons in the Adult Brain[M]. Yeshiva University, 2013.
This protocol only provides a guideline, and should be modified according to your specific needs.
1. Solution preparation
(1) Stock solution: Dissolve 5-BrdU solid in DMSO or deionized water to a final concentration of 10 mM and filter sterilize.
Note: After unused storage solution is aliquoted, store it in a dark place at -20℃ to avoid repeated freezing and thawing.
(2) Working solution: Before the formal experiment, dilute the stock solution with experimental buffer to the required working concentration and filter sterilize.
Note: Please adjust the working concentration of a specific application according to the actual situation or refer to the literature to set the gradient concentration by yourself to explore the best conditions. The working solution must be prepared and used immediately.
2. 5-BrdU in vitro labeling of cells
(1) Weigh 3 mg 5-BrdU and dissolve it in 1 ml deionized water until it is fully dissolved. Prepare 10 mM BrdU stock solution. Dilute the stock solution to 10 μM BrdU staining solution with cell culture medium at a ratio of 1:1000, and filter sterilize the staining solution with a 0.2 μm filter under sterile conditions.
(2) Aspirate the intracellular culture medium and replace with sterile 10µM BrdU staining solution, and incubate in a CO2 incubator at 37℃ for 1-24h.
(3) Aspirate the intracellular staining solution and wash twice with PBS, each time for at least 5s.
(4) Fix and permeabilize the cells according to the standard immunocytochemistry (ICC) operation process. However, before starting immunostaining, refer to the following DNA denaturation steps.
Note: The 5-BrdU incubation time depends on the cell division rate. Primary cells may take up to 24h, but fast-proliferating cell lines may only take 1h. The time to achieve the best signal-to-noise ratio needs to be determined by optimizing conditions.
3. 5-BrdU in vivo labeling
There are many methods for 5-BrdU in vivo labeling, and the most common one is intraperitoneal injection. The following are the steps for intraperitoneal injection. Other methods can refer to the literature or operate according to the inherent experience of the laboratory.
(1) Dissolve an appropriate amount of 5-BrdU in 1x PBS to prepare a 10 mg/ml stock solution, filter and sterilize, and freeze it in single-use aliquots for later use. Avoid repeated freezing and thawing.
(2) For mice, the commonly used injection concentration of 5-BrdU is 100 mg/kg. After labeling, treat the animals according to established laboratory procedures.
Note: 5-BrdU insertion into rapidly differentiating tissues such as the small intestine can be detected 30 minutes after injection. However, most tissues may require up to 24 hours to be detected. The appropriate treatment time and injection dose need to be optimized based on the tissue type.
4. DNA denaturation step
Cell sample:
(1) Incubate cells with 1-2.5M HCl at room temperature for 10 minutes to 1 hour. The actual HCl concentration and incubation time are optimized based on the experiment. If a shorter incubation time is used, incubation at 37°C may be more effective than incubation at room temperature.
(2) Optional step: Remove HCl and neutralize with 0.1M sodium borate buffer, pH 8.5 at room temperature for 30 minutes.
(3) Wash with PBS three times, each time for at least 5 seconds. Continue with immunostaining according to the standard IHC procedure.
Tissue sections:
(1) Dewax paraffin sections first.
(2) Incubate sections with 1-2.5M HCl at room temperature for 30 minutes to 1 hour. The actual HCl concentration and incubation time are optimized according to the experiment. If a shorter incubation time is used, incubation at 37°C may be more effective than incubation at room temperature.
(3) Optional step: Remove HCl and neutralize sections with 0.1M sodium borate buffer, pH 8.5 at room temperature for 10 minutes.
(4) Wash with PBS three times, each time for at least 5 seconds. Continue with immunostaining according to the standard IHC procedure.
Note: 5-BrdU is a mutagen. Be sure to take precautions during the operation and avoid direct contact with the skin. In addition, wear a mask to avoid inhalation of dust. For your safety and health, please wear a lab coat and disposable gloves.
| Cas No. | 59-14-3 | SDF | |
| المرادفات | BrdU, Bromodeoxyuridine, Broxuridine, NSC 38297 | ||
| Chemical Name | 5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione | ||
| Canonical SMILES | BrC(C(N1)=O)=CN(C1=O)[C@@]2([H])C[C@@](O)([H])[C@@](O2)([H])CO | ||
| Formula | C9H11BrN2O5 | M.Wt | 307.1 |
| الذوبان | ≥ 15.35mg/mL in DMSO, ≥ 16.23 mg/mL in Water with ultrasonic, ≥ 11.56 mg/mL in 0.9% NS with ultrasonic | Storage | 4°C, protect from light |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
||
| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
|
1 mg | 5 mg | 10 mg |
| 1 mM | 3.2563 mL | 16.2813 mL | 32.5627 mL |
| 5 mM | 0.6513 mL | 3.2563 mL | 6.5125 mL |
| 10 mM | 0.3256 mL | 1.6281 mL | 3.2563 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
g
μL
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >99.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 20 reference(s) in Google Scholar.)
GLPBIO products are for RESEARCH USE ONLY. Please make sure your review or question is research based.
Required fields are marked with *