Bathophenanthroline Sulfonate-Based Colorimetric Assay: A Simple and Rapid Method for Quantitation of Non-Heme Iron in Mouse Liver Tissue
Published: April 30, 2023
Abstract
Source: Duarte, T. L., Neves, J. V. Measurement of Tissue Non-Heme Iron Content using a Bathophenanthroline-Based Colorimetric Assay. J. Vis. Exp. (2022)
In this video, we describe a colorimetric-based method to determine non-heme iron content in mouse liver tissue. Bathophenanthroline sulfonate is used as the chromogenic substrate in the presence of a reducing agent, which converts ferric iron to chelatable ferrous iron.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board 1. Solution preparation NOTE: Handle and prepare all reagents and solutions with iron-free glassware or disposable plasticware. Do not allow metallic laboratory materials (e.g., stainless steel spatulas) to come in contact with any reagent or solution, due to the risk of iron contamination. Ma…
Disclosures
The authors have nothing to disclose.
Materials
| 96 well UV transparent plate | Sarstedt | 82.1581.001 | |
| Analytical balance | Kern | ABJ 220-4M | |
| Anhydrous sodium acetate | Merck | 106268 | |
| Bathophenanthroline sulfonate (4,7-Diphenyl-1,10-phenantroline dissulfonic acid) | Sigma-Aldrich | B1375 | |
| C57BL/6 mice (Mus musculus) | Charles River Laboratories | ||
| Carbonyl iron powder, ≥99.5% | Sigma-Aldrich | 44890 | |
| Disposable cuvettes in polymethyl methacrylate (PMMA) | VWR | 634-0678P | |
| Double distilled, sterile water | B. Braun | 0082479E | |
| Fluorescence microplate reader | BioTek Instruments | FLx800 | |
| Hydrochloric acid, 37% | Sigma-Aldrich | 258148 | |
| Microwave digestion oven and white teflon cups | CEM | MDS-2000 | |
| Nitric acid | Fisher Scientific | 15687290 | |
| Oven | Binder | ED115 | |
| Rodent chow | Harlan Laboratories | 2014S | Teklad Global 14% Protein Rodent Maintenance Diet containing 175 mg/kg iron |
| Sea bass (Dicentrarchus labrax) | Sonrionansa | ||
| Sea bass feed | Skretting | L-2 Alterna 1P | |
| Single beam UV-Vis spectrophotometer | Shimadzu | UV mini 1240 | |
| Thioglycolic acid | Merck | 100700 | |
| Trichloroacetic acid | Merck | 100807 |
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Cite This Article
Bathophenanthroline Sulfonate-Based Colorimetric Assay: A Simple and Rapid Method for Quantitation of Non-Heme Iron in Mouse Liver Tissue. J. Vis. Exp. (Pending Publication), e21159, doi: (2023).