3,3′,5,5′-Tetramethylbenzidine dihydrochloride, CAS No.64285-73-0

CAS: 64285-73-0 Cat. No.: T755451 Molecular Weight: 313.27 EC Number: 264-769-6 PubChem CID: 174033

GRADE & PURITY tablet, 1 mg substrate per tablet

Overview

3,3′,5,5′-Tetramethylbenzidine dihydrochloride has been used as a substrate for the

• anti-mouse IgG (? specific) horseradish peroxidase-conjugated antibody in the ELISA of mice sera
• anti-mouse peroxidase antibody in the protein tyrosine phosphatase assay of naive and effector T cell lysate
• in enzymatic hydroperoxide (EH) assay and in prooxidants–antioxidants balance (PAB) assay of serum samples from diabetic patients

3,3′,5,5′-Tetramethylbenzidine (TMB) is a peroxidase substrate suitable for use in ELISA procedures. The substrate produces a soluble end product that is pale blue in color and can be read spectrophotometrically at 370 or 620-650 nm. The TMB reaction may be stopped with 2 M H₂SO₄ (resulting in a yellow color), and read at 450 nm.

Specifications

Product Name

3,3′,5,5′-Tetramethylbenzidine dihydrochloride, CAS No.64285-73-0

Specifications & Purity

tablet, 1 mg substrate per tablet

CAS

64285-73-0

Molecule Type

Small molecule

Storage and Shipping

Concentration

tablet, 1 mg substrate per tablet

Storage

Store at 2-8°C

Shipped In

Wet ice

Taxonomic Classification

Taxonomy Tree

• Kingdom Organic compounds
  • Superclass Benzenoids
    • Class Benzene and substituted derivatives
      • Subclass Biphenyls and derivatives

• Kingdom: Organic compounds
• Superclass: Benzenoids
• Class: Benzene and substituted derivatives
• Subclass: Biphenyls and derivatives
• Intermediate Tree Nodes: Not available
• Direct Parent: Benzidines
• Alternative Parents: m-Xylenes Aniline and substituted anilines Primary amines Organopnictogen compounds Hydrochlorides Hydrocarbon derivatives
• Molecular Framework: Aromatic homomonocyclic compounds
• Substituents: Benzidine - M-xylene - Xylene - Aniline or substituted anilines - Organic nitrogen compound - Organopnictogen compound - Hydrocarbon derivative - Hydrochloride - Primary amine - Organonitrogen compound - Amine - Aromatic homomonocyclic compound
• Description: This compound belongs to the class of organic compounds known as benzidines. These are organic compounds containing the benzidine skeleton, made up of a biphenyl ring system substituted at the 4- and 4'-positions with a unsubstituted amine group.
• External Descriptors: Not available

Data sources

1. Djoumbou Feunang Y, Eisner R, Knox C, Chepelev L, Hastings J, Owen G, Fahy E, Steinbeck C, Subramanian S, Bolton E, Greiner R, and Wishart DS. ClassyFire: Automated Chemical Classification With A Comprehensive, Computable Taxonomy. Journal of Cheminformatics, 2016, 8:61.

DOI: 10.1186/s13321-016-0174-y

References

1. Yuan Zhang, Dong Wang, Gaili Liu, Yongjia Xiang, Kun Wang, Shaomin Shuang, Chuan Dong.  (2024)  All-in-one high performance CuO@MnO2 nanozymes: Dual-signal channel 2-mercaptobenzothiazole sensing and device-free oxidative degradation.  SENSORS AND ACTUATORS B-CHEMICAL,      [PMID:] [10.1016/j.snb.2024.136195]

2. Qiang Li, Saisong Zhan, Xiaoqing Yang, Zhaowei Zhang, Ning Sun, Xiang Wang, Jingjing Kang, Rui Du, Xiaoqin Hong, Minghao Yue, Xiaomin Li, Yujing Tang, Guangming Liu, Yue Liu, Dingbin Liu.  (2024)  Choline Phosphate-Grafted Nanozymes as Universal Extracellular Vesicle Probes for Bladder Cancer Detection.  ACS Nano,      [PMID:38857428] [10.1021/acsnano.4c00280]

3. Wang Yandong, Li Haodong, Lin Junyang, Li Yutang, Zhang Keqin, Li Hui, Fu Qiang, Jiang Yanyan.  (2025)  Engineering nanozyme immunomodulator with magnetic targeting effect for cascade-enzyodynamic and ultrasound-reinforced metallo-immunotherapy in prostate carcinoma.  Nature Communications,  16  (1): (1-23).  [PMID:39987131] [10.1038/s41467-025-57190-1]

4. Meng Ding, Tian Meng, Lingjie Song, Jian Sun, Lei Wang, Jinghua Yin, Shifang Luan.  (2024)  Generating Reactive Oxygen Species by a Dual-Catalytic Metal–organic Framework-based Nanozyme against Drug-resistant Bacteria Infection.  CHEMISTRY OF MATERIALS,      [PMID:] [10.1021/acs.chemmater.4c00646]

5. Furong Chao, Chengliang Cao, Yin Xu, Yunjie Gu, Xinyu Qu, Qian Wang, Lu−Lu Qu, Yuxin Guo, Xiaochen Dong.  (2025)  Sprayable Hydrogel for pH-Responsive Nanozyme-Derived Bacteria-Infected Wound Healing.  ACS Applied Materials & Interfaces,      [PMID:39831592] [10.1021/acsami.4c18100]

6. Meiqi Chang, Lu Zhang, Tingting Zhang, Yanqiu Duan, Wei Feng, Shaoling Yang, Yu Chen, Zeyu Wang.  (2024)  Ultrasound-augmented enzyodynamic-Ca2+ overload synergetic tumor nanotherapy.  BIOMATERIALS,      [PMID:38432005] [10.1016/j.biomaterials.2024.122513]

7. Shiyue Wu, Chaoqin Han, Wensheng Fu, Xue Gong, Zhongwei Jiang, Yi Wang, Pu Zhang.  (2023)  Smartphone-enabled colorimetric determination of silver ions exploiting the peroxidase-mimetic behavior of bowl-shaped Pd3Pt nanocrystals.  MICROCHEMICAL JOURNAL,      [PMID:] [10.1016/j.microc.2023.109434]

8. Yuting Wang, Tong Li, Hui Wei.  (2023)  Determination of the Maximum Velocity of a Peroxidase-like Nanozyme.  ANALYTICAL CHEMISTRY,      [PMID:37341651] [10.1021/acs.analchem.3c01830]

9. Ran Tang, Xiaoyu Yuan, Zhi Jia, Fang Yang, Gang Ye, Jie Liu.  (2023)  Ruthenium Dioxide Nanoparticles Treat Alzheimer’s Disease by Inhibiting Oxidative Stress and Alleviating Neuroinflammation.  ACS Applied Nano Materials,      [PMID:] [10.1021/acsanm.3c01650]

10. Wenxiang Du, Weijian Chen, Jing Wang, Hongjie Zhang, Lei Song, Yuan Hu, Xiaopeng Ma.  (2023)  A dual-nanozyme-loaded black phosphorus multifunctional therapeutic platform for combined photothermal/photodynamic/starvation cancer therapy.  Journal of Materials Chemistry B,  11  (23): (5185-5194).  [PMID:37264903] [10.1039/D3TB00372H]

11. Yuyun Ye, Hongli Yu, Bohan Chen, Yifan Zhao, Bai Lv, Guanghe Xue, Yong Sun, Jie Cao.  (2023)  Engineering nanoenzymes integrating Iron-based metal organic frameworks with Pt nanoparticles for enhanced Photodynamic-Ferroptosis therapy.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:37178565] [10.1016/j.jcis.2023.05.003]

12. Zhijun Huang, Xiaoting Li, Yanting Feng, Yonghui Tian, Yixiang Duan.  (2023)  Sensitive visual detection of gram-positive bacteria by composite filter membrane combined with antibiotic modified nanoparticles.  SENSORS AND ACTUATORS B-CHEMICAL,      [PMID:] [10.1016/j.snb.2023.133817]

13. Shunmei Li, Jie Xiang, Fang Yang, Ruo Yuan, Yun Xiang.  (2023)  Aptamer/proximity hybridization-based label-free and highly sensitive colorimetric detection of methotrexate via polymerization/nicking recycling amplifications.  SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY,      [PMID:36965245] [10.1016/j.saa.2023.122633]

14. Qi Xu, Han Hu, Zhimin Mo, Tianyou Chen, Qianyuan He, Zushun Xu.  (2023)  A multifunctional nanotheranostic agent based on Lenvatinib for multimodal synergistic hepatocellular carcinoma therapy with remarkably enhanced efficacy.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:36746055] [10.1016/j.jcis.2023.01.144]

15. Li Zhang, Wendong Zhang, Yao Nie, Yi Wang, Pu Zhang.  (2023)  Covalent organic framework-supported ultrasmall Rh nanoparticles as peroxidase mimics for colorimetric sensing of cysteine.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:36669450] [10.1016/j.jcis.2023.01.020]

16. Zhenyu Ding, Zhe Li, Xiaoxue Zhao, Yanrong Miao, Zhenfeng Yuan, Yuanyuan Jiang, Yizhong Lu.  (2022)  Self-deposited ultrasmall Ru nanoparticles on carbon nitride with high peroxidase-mimicking activity for the colorimetric detection of alkaline phosphatase.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:36368217] [10.1016/j.jcis.2022.11.001]

17. Yutong Li, Xinhui Gu, Jiayin Zhao, Fengna Xi.  (2022)  Fabrication of a Ratiometric Fluorescence Sensor Based on Carbon Dots as Both Luminophores and Nanozymes for the Sensitive Detection of Hydrogen Peroxide.  MOLECULES,  27  (21): (7379).  [PMID:36364206] [10.3390/molecules27217379]

18. Jie Xiang, Junyi Zhang, Shunmei Li, Ruo Yuan, Yun Xiang.  (2022)  Aptamer-based and sensitive label-free colorimetric sensing of phenylalanine via cascaded signal amplifications.  ANALYTICA CHIMICA ACTA,      [PMID:36192064] [10.1016/j.aca.2022.340393]

19. Yuan Yuni, liu Wei, Liu Qi, He Qing, Hu Yuyang, Jiang Xinyu, Chen Xiaoqing.  (2022)  A dual-channel probe based on copper ion-mediated metal organic framework composite for colorimetric and ratiometric fluorescence monitoring of glyphosate degradation in soil and water.  MICROCHIMICA ACTA,  189  (10): (1-10).  [PMID:36066687] [10.1007/s00604-022-05473-5]

20. Peilei Liu, Penghao Ji, Liping Wang, Haiyan Guo, Minfeng Huo, Jianlin Shi.  (2022)  Concurrent antibiosis and anti-inflammation against bacterial pneumonia by zinc hexacyanoferrate nanocatalysts.  BIOMATERIALS,      [PMID:36088676] [10.1016/j.biomaterials.2022.121768]

21. Shuoying Huang, Wenqiang Lai, Bingqian Liu, Mingdi Xu, Junyang Zhuang, Dianping Tang, Youxiu Lin.  (2022)  Colorimetric and photothermal dual-mode immunoassay of aflatoxin B1 based on peroxidase-like activity of Pt supported on nitrogen-doped carbon.  SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY,      [PMID:36049298] [10.1016/j.saa.2022.121782]

22. Yaqian He, Mengxue Yang, Tao Wang, Liuyang Xu, Zhenhe Ma, Jian Mu, Desong Wang, Dawei Gao.  (2022)  Regulation of Interstitial Fluid Pressure for Improved Intratumor Drug Permeability and Nanozyme-Enhanced Chemocatalytic Therapy.  ACS Sustainable Chemistry & Engineering,      [PMID:] [10.1021/acssuschemeng.2c02129]

23. Xiaoyan Gan, Feng Qiu, Ruo Yuan, Yun Xiang.  (2022)  Biobarcode and sequence-responsive DNA nanocapsules for label-free and sensitive electrochemical detection of circulating tumor cells.  SENSORS AND ACTUATORS B-CHEMICAL,      [PMID:] [10.1016/j.snb.2022.132207]

24. Chang Ping Yang, Cong Yi Hu, Zhong Wei Jiang, Si Yu Xiao, Xiao Yan Wang, Cheng Zhi Huang, Yuan Fang Li, Shu Jun Zhen.  (2022)  Facile synthesis of porphyrin-MOFs with high photo-Fenton activity to efficiently degrade ciprofloxacin.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:35533483] [10.1016/j.jcis.2022.04.104]

25. Song Meng, Zhihao Yao, Jiawei Liu, Erjing Wang, Cao Li, Bingbing Jiang, Ziqiang Xu.  (2022)  Carbon dots capped cerium oxide nanoparticles for highly efficient removal and sensitive detection of fluoride.  JOURNAL OF HAZARDOUS MATERIALS,      [PMID:35472541] [10.1016/j.jhazmat.2022.128976]

26. Tao Chen, Yuanhong Min, Xiao Yang, Hao Gong, Xiaoying Tian, Li Liu, Yanhua Hou, Wensheng Fu.  (2022)  Designing CoS1.035 Nanoparticles Anchored on N-Doped Carbon Dodecahedron as Dual-Enzyme Mimics for the Colorimetric Detection of H2O2 and Glutathione.  ACS Omega,      [PMID:35415371] [10.1021/acsomega.1c07264]

27. Mengying Luo, Mufang Li, Ying Lu, Ming Xia, Qinghua Zhao, Dong Wang.  (2022)  In-situ growth of multienzyme-inorganic hybrid nanoflowers on PVA-co-PE nanofibrous strip for colorimetric biosensor.  COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS,      [PMID:] [10.1016/j.colsurfa.2022.128419]

28. Yufeng Liu, Xiaoyu Wang, Quan Wang, Yihong Zhang, Quanyi Liu, Shujie Liu, Sirong Li, Yan Du, Hui Wei.  (2021)  Structurally Engineered Light-Responsive Nanozymes for Enhanced Substrate Specificity.  ANALYTICAL CHEMISTRY,      [PMID:34738799] [10.1021/acs.analchem.1c03610]

29. Pu Zhang, Wanqiang Xia, Ping Deng, Yuanhong Min, Juan Tan, Yi Wang, Wensheng Fu.  (2021)  Fe(III)-mediated reversible catalytic activity of MoS2 nanozymes for bisphosphonate drug sensing.  COLLOIDS AND SURFACES B-BIOINTERFACES,      [PMID:34218013] [10.1016/j.colsurfb.2021.111953]

30. Liu-Chuang Zhao, Zhen-Ni Chen, Pengcheng Wang, Qi Mou, Sheng-Hong Wu, Jian-Jun Sun, Jian-Hui Fang, Yimin Fang.  (2021)  Monitoring colorless electroactive chemicals in complex background based on electrochemical difference absorption spectroscopy with twin flow cells.  ANALYTICA CHIMICA ACTA,      [PMID:33992217] [10.1016/j.aca.2021.338521]

31. Juan Tan, Shiyue Wu, Qingqing Cai, Yi Wang, Pu Zhang.  (2021)  Reversible regulation of enzyme-like activity of molybdenum disulfide quantum dots for colorimetric pharmaceutical analysis.  Journal of Pharmaceutical Analysis,      [PMID:35573882] [10.1016/j.jpha.2021.03.010]

32. Xiaoyu Wang, Xiaoqian Jiang, Hui Wei.  (2020)  Phosphate-responsive 2D-metal–organic-framework-nanozymes for colorimetric detection of alkaline phosphatase.  Journal of Materials Chemistry B,  8  (31): (6905-6911).  [PMID:32424386] [10.1039/C9TB02542A]

33. Xin Yan, Chang Xia, Bin Chen, Yuan Fang Li, Peng Fei Gao, Cheng Zhi Huang.  (2019)  Enzyme Activity Triggered Blocking of Plasmon Resonance Energy Transfer for Highly Selective Detection of Acid Phosphatase.  ANALYTICAL CHEMISTRY,      [PMID:31850751] [10.1021/acs.analchem.9b04685]

34. Yahuan Wang, Chengyu Lin, Zhiwei Wang, Zhimin Chen, Jiafu Chen, Yong Chen, Shaohua Liu, Jianwei Fu.  (2019)  Magnetic hollow poly(cyclotriphosphazene-co-4,4′-sulfonyldiphenol)-Fe3O4 hybrid nanocapsules for adsorbing Safranine T and catalytic oxidation of 3,3′,5,5′-tetramethylbenzidine.  JOURNAL OF COLLOID AND INTERFACE SCIENCE,      [PMID:31454620] [10.1016/j.jcis.2019.08.073]

35. Cao Wen, Lin Junshu, Muhammad Faheem, Wang Quan, Wang Xiaoyu, Lou Zhangping, Wei Hui.  (2019)  Porous Ruthenium Selenide Nanoparticle as a Peroxidase Mimic for Glucose Bioassay.  Journal of Analysis and Testing,  3  (3): (253-259).  [PMID:] [10.1007/s41664-019-00104-0]

36. Yufeng Liu, Min Zhou, Wen Cao, Xiaoyu Wang, Quan Wang, Sirong Li, Hui Wei.  (2019)  Light-Responsive Metal–Organic Framework as an Oxidase Mimic for Cellular Glutathione Detection.  ANALYTICAL CHEMISTRY,      [PMID:31148448] [10.1021/acs.analchem.9b00512]

37. Nuanfei Zhu, Lantian Gu, Jin Wang, Xuesong Li, Guoxi Liang, Jinhui Zhou, Zhen Zhang.  (2019)  Novel and Sensitive Chemiluminescence Sensors Based on 2D-MOF Nanosheets for One-Step Detection of Glucose in Human Urine.  Journal of Physical Chemistry C,      [PMID:] [10.1021/acs.jpcc.9b00671]

38. Wang Yi-Wei, Liu Qiao, Wang Lixing, Tang Shurong, Yang Huang-Hao, Song Hongbo.  (2018)  A colorimetric mercury(II) assay based on the Hg(II)-stimulated peroxidase mimicking activity of a nanocomposite prepared from graphitic carbon nitride and gold nanoparticles.  MICROCHIMICA ACTA,  186  (1): (1-8).  [PMID:30535761] [10.1007/s00604-018-3137-3]

39. Li Qin, Xiaoyu Wang, Yufeng Liu, Hui Wei.  (2018)  2D-Metal–Organic-Framework-Nanozyme Sensor Arrays for Probing Phosphates and Their Enzymatic Hydrolysis.  ANALYTICAL CHEMISTRY,      [PMID:30044077] [10.1021/acs.analchem.8b02428]

40. Li-Juan Wang, Li-Ping Jia, Rong-Na Ma, Wen-Li Jia, Huai-Sheng Wang.  (2017)  A colorimetric assay for Hg2+ detection based on Hg2+-induced hybridization chain reactions.  Analytical Methods,  9  (35): (5121-5126).  [PMID:] [10.1039/C7AY01231D]

41. Liping Jia, Shanshan Shi, Rongna Ma, Wenli Jia, Huaisheng Wang.  (2016)  Highly sensitive electrochemical biosensor based on nonlinear hybridization chain reaction for DNA detection.  BIOSENSORS & BIOELECTRONICS,      [PMID:26872213] [10.1016/j.bios.2016.02.007]

42. Qun Wei Shu, Chun Mei Li, Peng Fei Gao, Ming Xuan Gao, Cheng Zhi Huang.  (2015)  Porous hollow CuS nanospheres with prominent peroxidase-like activity prepared in large scale by a one-pot controllable hydrothermal step.  RSC Advances,  5  (23): (17458-17465).  [PMID:] [10.1039/C4RA14609C]

43. Peng Zhao, Xuhui Xia, Yiyao Luo, Zirui Yuan, Jiaxi Deng, Huibo Luo, Xiaogang Luo, Danqun Huo, Changjun Hou.  (2024)  A porphyrin-modified CoMoO4 nanosensor array for the detection of crude baijiu.  Analytical Methods,      [PMID:39364608] [10.1039/D4AY01082E]

44. Hua-Ying Hou, Xu Chu, Guo-Dong Cheng, Zhuo-Yuan Li, Yi Liu, Shu- Lan Li.  (2025)  A Sandwich-like metabolic regulator activated γδ T cells for dual immunotherapy.  JOURNAL OF CONTROLLED RELEASE,      [PMID:41319962] [10.1016/j.jconrel.2025.114473]

45. Liuyi Huang, Xinhui Gu, Fengna Xi.  (2026)  Facile Aptasensor with enhanced electrochemiluminescence by nanozyme in nanoporous silica film for highly sensitive detection of carbohydrate antigen 15–3.  JOURNAL OF ELECTROANALYTICAL CHEMISTRY,      [PMID:] [10.1016/j.jelechem.2026.119896]

46. Yuanyuan Miao, Chuang Wang, Yue Peng, Xinyuan Sun, Zhaokun Zheng, Qianlei Zhang, Wenting Cheng, Jinlong Li.  (2026)  PAM-assembled CRISPR-Cas12a activation-based fluorescent and colorimetric dual-modal biosensor for detecting prostate cancer exosomes.  BIOSENSORS & BIOELECTRONICS,      [PMID:] [10.1016/j.bios.2026.118635]