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> DTNB [69-78-3]

DTNB [69-78-3]

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Cat# HY-15915-1g

Size : 1g

Brand : MedChemExpress

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Description

DTNB (Ellman’s Reagent) is a chemical used to quantify the number or concentration of thiol groups[1].

In Vitro

DTNB reacts with the free sulfhydryl side chain of cysteine to form an S-S bond between the protein and a thionitrobenzoic acid (TNB) residue[1].
The main advantage of DTNB over alternative reagents (e.g., N-ethylmaleimide or iodoacetamide) is in the selectivity of this reagent and in the ability to follow the course of the reaction spectrophotometrically[1].
Modification of Enzyme with DTNB:
Modification of the SH groups of the enzyme is carried out by reacting 10 μL of 10 mM DTNB solution (about a 20-fold molar excess) at room temperature with 0.6 mL of enzyme solution (0.807 mg/mL) in 80 mM phosphate buffer, pH 8.0, containing 1 mM EDTA, which has been dialyzed previously against the same buffer solution for 24 h. The number of SH groups is estimated from the increase of absorbance at 410 nm using a molar extinction coefficient of 13,600 M-1cm-1 for thionitrobenzoate ions liberated.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

DTNB Related Antibodies

Solvent & Solubility
In Vitro: 

DMSO : 100 mg/mL (252.30 mM; ultrasonic and warming and heat to 60°C; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)

H2O : < 0.1 mg/mL (insoluble)

Preparing
Stock Solutions
Concentration Solvent Mass 1 mg 5 mg 10 mg
1 mM 2.5230 mL 12.6151 mL 25.2302 mL
5 mM 0.5046 mL 2.5230 mL 5.0460 mL
View the Complete Stock Solution Preparation Table

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.

Select the appropriate dissolution method based on your experimental animal and administration route.

For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for in vivo experiments, it is recommended to prepare freshly and use it on the same day.
The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.

  • Protocol 1

    Add each solvent one by one:  10% DMSO    40% PEG300    5% Tween-80    45% Saline

    Solubility: ≥ 2.08 mg/mL (5.25 mM); Clear solution

    This protocol yields a clear solution of ≥ 2.08 mg/mL (saturation unknown).

    Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (20.8 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.

    Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
  • Protocol 2

    Add each solvent one by one:  10% DMSO    90% (20% SBE-β-CD in Saline)

    Solubility: ≥ 2.08 mg/mL (5.25 mM); Clear solution

    This protocol yields a clear solution of ≥ 2.08 mg/mL (saturation unknown).

    Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (20.8 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.

    Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
Purity & Documentation
References

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