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Hoechst 33258 staining solution (ready to use) - 5ug/ml in H2O, high purity , CAS No.23491-45-4

    Grade & Purity:
  • 5ug/ml in H2O
In stock
Item Number
H598341
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H598341-10ml
10ml
Available within 8-12 weeks(?)
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$73.90
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Cell staining (27) Nuclear dye (3)

Basic Description

Specifications & Purity 5ug/ml in H2O
Storage Temp Protected from light,Store at -20°C
Shipped In
Ice chest + Ice pads
This product requires cold chain shipping. Ground and other economy services are not available.
Product Description

Hoechst 33258, also known as bis benzimide h 33258 or hoe 33258, is a non embedding bright blfluorescent dye. Dyes have weak fluorescence in solution, and their fluorescence becomes bright after binding with DNA at the minor groove in the DNA poly at sequence rich region in living cells. Therefore, such dyes are also known as DNA probes. Because of the low background, the stained cells do not need washing steps, and the staining is very stable, non-toxic to live cells, and can last for several days or longer after combined with DNA staining. Hoechst 33258 has higher solubility in water than Hoechst 33342, but both dyes have high cell membrane permeability and are widely used for apoptosis detection. After staining, it can be observed by fluorescence microscope or detected by flow cytometry. Take adherent cells (96 well plate) as an example, 100 per well μ L staining solution, 10 ml can be used for the staining of 100 wells.

Product parameters:

Ex/em (bound DNA) = 352/461 nm; Ex/Em (unbound DNA) = 346/460 nm

Matters needing attention:

1. please centrifuge the product to the bottom of the tube immediately before use, and then conduct subsequent experiments. 

2. if you need to adjust the concentration, please select H598341 and configure the appropriate working fluid concentration by yourself. 

3. fluorescent dyes have quenching problems. It is recommended to observe immediately after staining live cells or tissues. 

4. for your safety and health, please wear experimental clothes and disposable gloves.

Scope of application:

Nuclear staining

Experimental steps:
1. For fixed cells or tissues
(1) For cell or tissue samples, wash and remove the fixative appropriately after fixation. If immunofluorescence staining is required, first perform immunofluorescence staining, and then follow the subsequent steps to perform Hoechst 33258 staining.
(2) For adherent cells or tissue sections, add a small amount of Hoechst 33258 working solution and cover the sample. For suspended cells, add at least three times the volume of the sample to be tested and mix well. Leave at room temperature for 3-5 minutes.
(3) Remove Hoechst 33258 staining solution and wash 2-3 times with TBST, PBS, or physiological saline for 3-5 minutes each time.
Note: The cleaning steps are optional but not necessary and do not affect dyeing after cleaning.
(4) Observe directly under a fluorescence microscope or observe under a fluorescence microscope after sealing. When a cell undergoes apoptosis, the nucleus of the apoptotic cell can be seen to be densely stained or fragmented and densely stained.
2. For live cells or tissues
(1) Add an appropriate amount of Hoechst 33258 working solution and fully cover the sample to be stained. Typically, 1 mL of staining solution is required per well for a six well plate, and 100 mL is required per well for a 96 well plate μ L's staining solution.
(2) Incubate at room temperature in dark for 10-30 minutes.
(3) Discard the staining solution, wash 2-3 times with PBS, then add 50 μ Perform micrographs using PBS.
Note: The cleaning steps are optional but not necessary and do not affect dyeing after cleaning.

Taxonomic Classification

Taxonomy Tree

Kingdom Organic compounds
Superclass Organoheterocyclic compounds
Class Benzimidazoles
Subclass Phenylbenzimidazoles
Intermediate Tree Nodes Not available
Direct Parent Phenylbenzimidazoles
Alternative Parents N-arylpiperazines  Phenylimidazoles  Dialkylarylamines  N-methylpiperazines  1-hydroxy-2-unsubstituted benzenoids  Benzene and substituted derivatives  Heteroaromatic compounds  Trialkylamines  Azacyclic compounds  Organooxygen compounds  Hydrochlorides  Hydrocarbon derivatives  
Molecular Framework Aromatic heteropolycyclic compounds
Substituents Phenylbenzimidazole - N-arylpiperazine - 2-phenylimidazole - Dialkylarylamine - Tertiary aliphatic/aromatic amine - 1-hydroxy-2-unsubstituted benzenoid - Phenol - N-alkylpiperazine - N-methylpiperazine - Monocyclic benzene moiety - 1,4-diazinane - Benzenoid - Piperazine - Azole - Heteroaromatic compound - Imidazole - Tertiary aliphatic amine - Tertiary amine - Azacycle - Hydrochloride - Organooxygen compound - Organonitrogen compound - Hydrocarbon derivative - Organic oxygen compound - Organic nitrogen compound - Amine - Aromatic heteropolycyclic compound
Description This compound belongs to the class of organic compounds known as phenylbenzimidazoles. These are compounds containing a phenylbenzimidazole skeleton, which consists of a benzimidazole moiety where its imidazole ring is attached to a phenyl group.
External Descriptors Not available

Associated Targets(Human)

TOP1 Tclin DNA topoisomerase I (7553 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
TOP2A Tclin DNA topoisomerase II alpha (6317 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
DU-145 (51482 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
MCF7 (126967 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
A549 (127892 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
KG-1 (867 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
HeLa (62764 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID

Associated Targets(non-human)

Pseudomonas aeruginosa (123386 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Enterobacter cloacae (7976 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Klebsiella pneumoniae (43867 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Shigella flexneri (1836 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Staphylococcus aureus (210822 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Staphylococcus epidermidis (22802 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Acinetobacter baumannii (41033 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Citrobacter freundii (1864 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Escherichia coli (133304 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Enterococcus faecium (13803 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Enterococcus faecalis (29875 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID
Calf thymus DNA (4845 Activities)
Activity Type Relation Activity value Units Action Type Journal PubMed Id doi Assay Aladdin ID

Mechanisms of Action

Mechanism of Action Action Type target ID Target Name Target Type Target Organism Binding Site Name References

Names and Identifiers

IUPAC Name 4-[6-[6-(4-methylpiperazin-1-yl)-1H-benzimidazol-2-yl]-1H-benzimidazol-2-yl]phenol;trihydrochloride
INCHI InChI=1S/C25H24N6O.3ClH/c1-30-10-12-31(13-11-30)18-5-9-21-23(15-18)29-25(27-21)17-4-8-20-22(14-17)28-24(26-20)16-2-6-19(32)7-3-16;;;/h2-9,14-15,32H,10-13H2,1H3,(H,26,28)(H,27,29);3*1H
InChIKey SMNPLAKEGAEPJD-UHFFFAOYSA-N
Smiles CN1CCN(CC1)C2=CC3=C(C=C2)N=C(N3)C4=CC5=C(C=C4)N=C(N5)C6=CC=C(C=C6)O.Cl.Cl.Cl
Isomeric SMILES CN1CCN(CC1)C2=CC3=C(C=C2)N=C(N3)C4=CC5=C(C=C4)N=C(N5)C6=CC=C(C=C6)O.Cl.Cl.Cl
WGK Germany 3
RTECS SM1140500
PubChem CID 31953
Molecular Weight 533.90
Beilstein 4088183

Certificates(CoA,COO,BSE/TSE and Analysis Chart)

C of A & Other Certificates(BSE/TSE, COO):
Analytical Chart:

Find and download the COA for your product by matching the lot number on the packaging.

1 results found

Lot Number Certificate Type Date Item
A2420226 Certificate of Analysis Jan 16, 2024 H598341

Chemical and Physical Properties

Melt Point(°C) 314°C
Molecular Weight 533.900 g/mol
XLogP3
Hydrogen Bond Donor Count 6
Hydrogen Bond Acceptor Count 5
Rotatable Bond Count 3
Exact Mass 532.131 Da
Monoisotopic Mass 532.131 Da
Topological Polar Surface Area 84.100 Ų
Heavy Atom Count 35
Formal Charge 0
Complexity 634.000
Isotope Atom Count 0
Defined Atom Stereocenter Count 0
Undefined Atom Stereocenter Count 0
Defined Bond Stereocenter Count 0
Undefined Bond Stereocenter Count 0
The total count of all stereochemical bonds 0
Covalently-Bonded Unit Count 4

Citations of This Product

1. Weiyuan Ma, Xiuwen Zhang, Le Zhuang.  (2023)  Exogenous Hydrogen Sulfide Induces A375 Melanoma Cell Apoptosis Through Overactivation of the Unfolded Protein Response.  Clinical Cosmetic and Investigational Dermatology,     
2. Yingying Zhang, Tiange Wang, Yinping Zhuang, Tiandi He, Xiaoli Wu, Lin Su, Jun Kang, Jin Chang, Hanjie Wang.  (2021)  Sodium Alginate Hydrogel-Mediated Cancer Immunotherapy for Postoperative In Situ Recurrence and Metastasis.  ACS Biomaterials Science & Engineering,  (12): (5717–5726). 
3. Yunran Zhang, Haiping Zhou, Zhe Zhang, Yijun Zhu, Tao Wang, Liguo Yu, Hongrui Song.  (2021)  Redox/NIR dual-responsive PEG-betulinic acid/pluronic-cypate prodrug micelles for chemophotothermal therapy.  COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS,  609  (125662). 
4. Chenglong Ma, Wei Sun, Waqas Qamar Zaman, Zhenhua Zhou, Hao Zhang, Qicheng Shen, Limei Cao, Ji Yang.  (2020)  Lanthanides Regulated the Amorphization–Crystallization of IrO2 for Outstanding OER Performance.  ACS Applied Materials & Interfaces,  12  (31): (34980–34989). 
5. Zhang Tao, Jiang Zhenqi, Chen Libin, Pan Chunshu, Sun Shan, Liu Chuang, Li Zihou, Ren Wenzhi, Wu Aiguo, Huang Pintong.  (2020)  PCN-Fe(III)-PTX nanoparticles for MRI guided high efficiency chemo-photodynamic therapy in pancreatic cancer through alleviating tumor hypoxia.  Nano Research,  13  (1): (273-281). 
6. Jun Dai, Yinghao Li, Zi Long, Ruming Jiang, Zeyan Zhuang, Zhiming Wang, Zujin Zhao, Xiaoding Lou, Fan Xia, Ben Zhong Tang.  (2020)  Efficient Near-Infrared Photosensitizer with Aggregation-Induced Emission for Imaging-Guided Photodynamic Therapy in Multiple Xenograft Tumor Models.  ACS Nano,  14  (1): (854–866). 
7. Tao Zhang, Zhenqi Jiang, Ting Xve, Shan Sun, Juan Li, Wenzhi Ren, Aiguo Wu, Pintong Huang.  (2019)  One-pot synthesis of hollow PDA@DOX nanoparticles for ultrasound imaging and chemo-thermal therapy in breast cancer.  Nanoscale,  11  (45): (21759-21766). 
8. Hailing Yang, Zhenjie Liu, Yonglong Song, Changjiang Hu.  (2019)  Hyaluronic acid-functionalized bilosomes for targeted delivery of tripterine to inflamed area with enhancive therapy on arthritis.  DRUG DELIVERY,  26  (1): (820-830). 
9. Zigui Wang, Gaizhen Kuang, Zhiqiang Yu, Aimin Li, Dongfang Zhou, Yubin Huang.  (2019)  Light-activatable dual prodrug polymer nanoparticle for precise synergistic chemotherapy guided by drug-mediated computed tomography imaging.  Acta Biomaterialia,  94  (459). 
10. Zhao Shuai, Sun Shan, Jiang Kai, Wang Yuhui, Liu Yu, Wu Song, Li Zhongjun, Shu Qinghai, Lin Hengwei.  (2019)  In Situ Synthesis of Fluorescent Mesoporous Silica–Carbon Dot Nanohybrids Featuring Folate Receptor-Overexpressing Cancer Cell Targeting and Drug Delivery.  Nano-Micro Letters,  11  (1): (1-13). 
11. Rui Wang, Xing Hu, Junhui Pan, Guowen Zhang, Deming Gong.  (2019)  Interaction of isoeugenol with calf thymus DNA and its protective effect on DNA oxidative damage.  JOURNAL OF MOLECULAR LIQUIDS,  282  (356). 
12. Jie Li, Suqin Shen, Fei Kong, Ting Jiang, Cui Tang, Chunhua Yin.  (2018)  Effects of pore size on in vitro and in vivo anticancer efficacies of mesoporous silica nanoparticles.  RSC Advances,  (43): (24633-24640). 
13. Jie-Hua Shi, Yan-Yue Lou, Kai-Li Zhou, Dong-Qi Pan.  (2018)  Exploration of intermolecular interaction of calf thymus DNA with sulfosulfuron using multi-spectroscopic and molecular docking techniques.  SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY,  204  (209). 
14. Ying Wang, Yating Zhao, Yu Cui, Qinfu Zhao, Qiang Zhang, Sara Musetti, Karina A Kinghorn, Siling Wang.  (2018)  Overcoming multiple gastrointestinal barriers by bilayer modified hollow mesoporous silica nanocarriers.  Acta Biomaterialia,  65  (405). 
15. Huanan Wang, Wenqing Xu.  (2017)  Mito-methyl coumarin, a novel mitochondria-targeted drug with great antitumor potential was synthesized.  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS,  489  (1). 
16. Hongxia Liu, Shuqin Wu, Jingmou Yu, Dun Fan, Jin Ren, Lei Zhang, Jianguo Zhao.  (2017)  Reduction-sensitive micelles self-assembled from amphiphilic chondroitin sulfate A-deoxycholic acid conjugate for triggered release of doxorubicin.  Materials Science & Engineering C-Materials for Biological Applications,  75  (55). 
17. Huanan Wang, Ming Yao, Wenqing Xu.  (2016)  The antitumor effects of mitochondria-targeted 6-(nicotinamide) methyl coumarin.  Open Life Sciences,  11  (1): (542-551). 
18. Haibo Mu, Qianjin Liu, Hong Niu, Dongdong Wang, Jiangjiang Tang, Jinyou Duan.  (2016)  Autophagy promotes DNA–protein crosslink clearance.  MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS,  797  (21). 
19. Li Jingchao, Wang Tingting, Shi Yuanfei, Ye Zichen, Zhang Xun, Ming Jiang, Zhang Yafei, Hu Xinyan, Li Yun, Zhang Dongsheng, Xu Qianhe, Yang Jun, Chen Xiaolan, Liu Nian, Su Xinhui.  (2024)  A continuously efficient O2-supplying strategy for long-term modulation of hypoxic tumor microenvironment to enhance long-acting radionuclides internal therapy.  JOURNAL OF NANOBIOTECHNOLOGY,  22  (1): (1-15). 
20. Yanghui Chen, Yuan Li, Junliang Luo, Zixin Li, Yu Huang, Jia Cai, Dongneng Jiang, Defeng Zhang, Jichang Jian, Jun Qiang, Bei Wang.  (2024)  A novel study of brain microvascular endothelial cells induced by astrocyte conditioned medium for constructing blood brain barrier model in vitro: A promising tool for meningitis of teleost.  FISH & SHELLFISH IMMUNOLOGY,  146  (109401). 
21. Ouyang Boshu, Shan Caihua, Shen Shun, Dai Xinnan, Chen Qingwang, Su Xiaomin, Cao Yongbin, Qin Xifeng, He Ying, Wang Siyu, Xu Ruizhe, Hu Ruining, Shi Leming, Lu Tun, Yang Wuli, Peng Shaojun, Zhang Jun, Wang Jianxin, Li Dongsheng, Pang Zhiqing.  (2024)  AI-powered omics-based drug pair discovery for pyroptosis therapy targeting triple-negative breast cancer.  Nature Communications,  15  (1): (1-20). 
22. Zhang Hongmei, Yang Yunxia, Wang Yanqing.  (2025)  Comprehensive Investigations About the Binding Interactions of Sudan Dyes with DNA by Spectroscopy and Docking Methods.  JOURNAL OF FLUORESCENCE,    (1-11). 
23. Ziwei Yan, Hui Zhang, Jinghao Chen, Qingqing Xu, Shuaipeng Feng, Qinfu Zhao, Siling Wang.  (2024)  Cu(II)-doped mesoporous polydopamine as biodegradable nanoplatforms for photothermal-enhanced multi-mode anti-tumor therapy.  COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS,  685  (133258). 
24. Jingmou Yu, Yifei Zhang, Meilin Xu, Dengzhao Jiang, Wenbo Liu, Hongguang Jin, Pu Chen, Jing Xu, Lei Zhang.  (2024)  Innovative gelatin-based micelles with AS1411 aptamer targeting and reduction responsiveness for doxorubicin delivery in tumor therapy.  BIOMEDICINE & PHARMACOTHERAPY,  174  (116446). 
25. Xiaowen Liu, Huamei Cai, Liang Peng, Hongli Ma, Yun Yan, Weixia Li, Jing Zhao.  (2024)  Microglial Nrf2/HO-1 signaling gates remifentanil-induced hyperalgesia via suppressing TRPV4-mediated M1 polarization.  FREE RADICAL BIOLOGY AND MEDICINE,     
26. CaoXiaona , LiJie , RenJinliang , PengJiajin , ZhongRuyue , HeJiahao , XuTing , YuZhenhua , JinHuawei , HaoSiqi , LiuRuiwei , XuBingzhe.  (2024)  Minimally-invasive implantable device enhances brain cancer suppression.  EMBO Molecular Medicine,     
27. Hao Liu, Ke Huang, Hao Zhang, Xiaohui Liu, Hui Jiang, Xuemei Wang.  (2024)  Photo-Driven In Situ Solidification of Whole Cells through Inhibition of Trogocytosis for Immunotherapy.  Research,  (2024 Feb;7). 
28. Junhong Ling, Yingying Cai, Haozhan Feng, Zhen Liu, Xiao-kun Ouyang.  (2024)  Polydopamine-Modified Copper Coordination Mesoporous Silica Nanoparticles Loaded with Disulfiram for Synergistic Chemo-Photothermal Therapy.  Pharmaceutics,  16  (4): (512). 
29. Zepeng Cui, Ruqian Ji, Jun Xie, Chao Wang, Jia Tian, Weian Zhang.  (2024)  Tumor Microenvironment-Triggered Self-Adaptive Polymeric Photosensitizers for Enhanced Photodynamic Therapy.  BIOMACROMOLECULES,  25  (4): (2302-2311). 

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