Tissue iron assay kit

Manufactured by Nanjing Jiancheng

Sourced in China

The Tissue Iron Assay Kit is a laboratory equipment used for the quantitative determination of iron content in biological samples such as tissues. It provides a reliable and accurate method to measure the iron levels in a wide range of sample types.

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26 protocols using tissue iron assay kit

Quantifying Murine Lung Iron Levels

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Paraffin-embedded murine right lung tissues were sectioned and stained with Prussian blue dye, and blue staining was evaluated by microscopy (BX51 Olympus, Japan). The iron concentration in the lungs was assessed with a Tissue Iron Assay Kit (Nanjing Jiancheng Bioengineering Institute, China).
We used FerroOrange (Dojindo, Chain) to detect intracellular Fe 2+ . Cells were incubated with 1 μmol/L FerroOrange for 30 min and visualized using fluorescence microscopy (BX51 Olympus, Japan).

Jiao Y., Yong C., Zhang R., Qi D, & Wang D. (2022). Hepcidin Alleviates LPS-Induced ARDS by Regulating the Ferritin-Mediated Suppression of Ferroptosis. Shock (Augusta, Ga.), 57(6).

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Iron and ROS Quantification in Spinal Cord and OPCs

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Spinal cord samples (n=8) or primary OPCs were collected and homogenized under ice-cold conditions. Iron levels were measured using the tissue iron assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China) in accordance with manufacturer's instruction. To determine iron concentration, the optical density (OD) value was measured at 520 nm using a spectrophotometer (Varioskan Flash, Thermo Scienti c, Waltham, MA, USA) and the iron concentration in all samples was then determined through comparing the OD of the samples to the standard curve.
Reactive oxygen species (ROS) measurement ROS levels were determined by a reactive oxygen species assay kit according to the manufacturer's speci cation (cat. no. S0033S, Beyotime Biotechnology, Beijing, China). Spinal cord samples (n=3) or primary OPCs were collected and homogenized under ice-cold conditions. Each sample was loaded 500 μl of 10 μM DCFH-DA and incubated in the dark at 37°C for 20 min. The loading buffer was replaced and washed three times to eliminate residual DCFH-DA. The samples were then immediately measured using a Flow cytometer (ACEA Biosciences Inc., San Diego, CA, USA) with an argon laser (488 nm).

Ge H., Xue X., Feng H., Yuan L., Wang L., Zou Y., Zhong J., Jiang Z., Shi J., Chen T., Hong S.Y., Feng H., & Hu S.J. (2021). Ferrostatin-1 Alleviates White Matter Injury via Decreasing Ferroptosis Following Spinal Cord Injury.

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Oxidative Stress Biomarkers in Liver

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The ROS was detected by Reactive Oxygen Species Fluorogenic Probe (BestBio; BB470513) and was observed with confocal microscopy. The frozen liver tissue (100 mg) was homogenized in 0.9 ml PBS. The lysate was centrifuged and the supernatant was collected. Hepatic MDA content, GSH content, iron content, TRF content was detected using the Malondialdehyde assay kit (Jiancheng, A003‐1‐2), Reduced glutathione assay kit (Jiancheng, A006‐2‐1), tissue iron assay kit (Jiancheng, A039‐2‐1), and Transferrin Assay Kit (Jiancheng, E028‐1‐1) following the manufacturer's instructions. The values for the levels of GSH in the liver tissues were measured using a microplate reader at the absorption wavelengths of 405nm. The values for the levels of MDA, iron, TRF in the liver tissues were measured using a spectrophotometer at the absorption wavelengths of 532, 520, and 340 nm.

Xing G., Meng L., Cao S., Liu S., Wu J., Li Q., Huang W, & Zhang L. (2022). PPARα alleviates iron overload‐induced ferroptosis in mouse liver. EMBO Reports, 23(8), e52280.

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Tissue Iron Level Quantification

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The collection of samples was first performed, and the spinal cord iron levels of rats in each group were determined using a tissue iron assay kit (Nanjing Jiancheng Biotechnology Co., Ltd.) according to the manufacturer's instructions. To determine the iron concentration, measure and read the optical density (OD) value at 520 nm using a spectrophotometer (Varioskan Flash; Thermo Scientific), then determine the iron concentration in all samples by contrasting the samples' OD with the standard curve.

Li D., Lu X., Xu G., Liu S., Gong Z., Lu F., Xia X., Jiang J., Wang H., Zou F, & Ma X. (2023). Dihydroorotate dehydrogenase regulates ferroptosis in neurons after spinal cord injury via the P53‐ALOX15 signaling pathway. CNS Neuroscience & Therapeutics, 29(7), 1923-1939.

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Quantifying Cellular Antioxidant and Lipid Peroxidation

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GSH is an important antioxidant, and the depletion of GSH can trigger ferroptosis [39 (link)]. Malondialdehyde (MDA) is an end-product of lipid peroxides, which is used to assess ferroptosis [40 (link)]. The cellular or tissue GSH, MDA content, tissue iron levels and serum iron levels were detected using the reduced glutathione (GSH) assay kit (A006-2-1, Nanjing Jiancheng Bioengineering Institute, Nanjing, China), the malondialdehyde (MDA) detection kit (S0131, Beyotime Biotechnology, Shanghai, China), the tissue iron assay kit (A039-2-1) and the serum iron assay kit (A039-1-1, Nanjing Jiancheng Bioengineering Institute, Nanjing, China) following the manufacturer’s instructions. The concentrations of GSH and MDA were determined by measuring the absorbance at 405 nm and 530 nm. The values for the levels of tissue and serum iron were examined at a wavelength of 520 nm.

Dong Q., Han Z., Gao M, & Tian L. (2024). FNDC5/irisin ameliorates bone loss of type 1 diabetes by suppressing endoplasmic reticulum stress‑mediated ferroptosis. Journal of Orthopaedic Surgery and Research, 19, 205.

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Hepatic Protein and Iron Quantification

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The liver proteins were extracted by the tissue homogenate method and detected using the BCA protein assay kit (PC0020; Solarbio, Beijing, China). Iron levels in the liver were measured using a tissue iron assay kit (Nanjing Jiancheng Bioengineering Institute). The optical density values were measured with a microplate reader.

Cheng H., Shi Y., Li X., Jin N., Zhang M., Liu Z., Liang Y, & Xie J. (2024). Human umbilical cord mesenchymal stem cells protect against ferroptosis in acute liver failure through the IGF1-hepcidin-FPN1 axis and inhibiting iron loading: Mesenchymal stem cells protect against ferroptosis in acute liver failure. Acta Biochimica et Biophysica Sinica, 56(2), 280-290.

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Oxidative Stress Markers in Spinal Cord Injury

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Iron accumulation in SCI was tested using the tissue iron assay kit (A039-2-1, Nanjing Jiancheng Bioengineering Institute). Malondialdehyde (MDA), as a natural product of lipid peroxidation, was quanti ed using Lipid Peroxidation MDA Assay Kit (S0131S, Beyotime). The protocols of iron and MDA concentration tests were performed according to the menufacturer's instructions and the absorbance was measured at 532 nm and 520 nm respectively. ROS assay ROS production was determined by chemiluninescence using the uorescent probe 2′, 7′-dichlorodihydro uorescein diacetate (DCFH-DA, WanleiBio). After anesthetization, rats were perfused transcardially with precooled PBS and the spinal cords were digested by pancreatin to obtain single-cell suspensions. Cells were incubated with DCFH-DA (10 μM) at 37℃ for 20 minutes. Later, the cells were observed and pictured via the inverted uorescence microscope (Axio Observer 3, Germany). The ROS uorescence intensity was calculated by software Image J.

Kang Y., Li Q., Zhu R., Li S., Xu X., Shi X, & Yin Z. (2022). Identification of Ferroptotic Genes in Spinal Cord Injury at Different Time Points: Bioinformatics and Experimental Validation. Molecular neurobiology, 59(9).

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Spinal Cord Iron Quantification

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Spinal cord samples were collected as described above and homogenized under ice-cold conditions. Iron levels of spinal cords at 2 days and 2 weeks post-SCI were detected by following the tissue iron assay kit instructions (Nanjing Jiancheng Bioengineering Institute, Nanjing, Jiangsu Province, China). All assays were performed in triplicate.

Hao J., Li B., Duan H.Q., Zhao C.X., Zhang Y., Sun C., Pan B., Liu C., Kong X.H., Yao X, & Feng S.Q. (2017). Mechanisms underlying the promotion of functional recovery by deferoxamine after spinal cord injury in rats. Neural Regeneration Research, 12(6), 959-968.

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Prostate Oxidative Stress Biomarkers

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Reactive oxygen species (ROS) in the prostate tissue was dyed with 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA; MedChemExpress, NJ, USA) probe and visualized under a fluorescence microscope (Olympus, Tokyo, Japan). To detect the oxidative stress degree, the content of lipid peroxidation product malondialdehyde (MDA) and the activities of antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) were measured with the thiobarbituric acid (TBA) method, xanthine oxidase (XO) method, and ammonium molybdate colorimetry method, respectively (all purchased from Nanjing Jiancheng Bioengineering Institute, Nanjing, China). The tissue iron assay kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China) was applied to determine the iron content of prostate lysates according to kit instruction. The protein concentration was detected to normalize the data.

Lin D., Zhang M., Luo C., Wei P., Cui K, & Chen Z. (2022). Targeting Ferroptosis Attenuates Inflammation, Fibrosis, and Mast Cell Activation in Chronic Prostatitis. Journal of Immunology Research, 2022, 6833867.

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Iron and ROS Quantification in Spinal Cord and OPCs

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Ge H., Xue X., Xian J., Yuan L., Wang L., Zou Y., Zhong J., Jiang Z., Shi J., Chen T., Su H., Feng H, & Hu S. (2022). Ferrostatin-1 Alleviates White Matter Injury Via Decreasing Ferroptosis Following Spinal Cord Injury. Molecular neurobiology, 59(1).

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