QuantiChrom™ Iron Assay Kit
Protocol SDS  Printed Protocols and SDSs are not provided with kits.

Application
For quantitative determination of iron ions Fe3+ and/or Fe2+ and evaluation of drug effects on iron metabolism.

Key Features
Sensitive and accurate. Linear detection range 27 μg/dL (4.8 μM) to 1,000 μg/dL (179 μM) iron in 96-well plate assay.

Simple and high-throughput. The procedure involves addition of a single working reagent and incubation for 40 min. Can be readily automated as a high-throughput assay for thousands of samples per day.

Improved reagent stability and versatility. The optimized formulation has greatly enhanced reagent and signal stability. Cuvette or 96-well plate assay.

Low interference in biological samples. No pretreatments are needed. Assays can be directly performed on serum samples.

Method
OD590nm

Samples
Biological (e.g. serum) and environmental samples

Species
All

Procedure
30 min

Size
250 tests

Detection Limit
27 μg/dL (4.8 μM)

Shelf Life
12 months

More Details
Iron level in blood is a reliable diagnostic indicator of various disease states. Increased levels of iron concentration in blood are associated with blood loss, increased destruction of red blood cells (e.g. hemorrhage) or decreased blood cell survival, acute hepatitis, certain sideroachrestic anemias, ingestion of iron-rich diets, defects in iron storage (e.g. pernicious anemia). Decreased levels of blood iron may result from insufficient iron ingestion from diets, chronic blood loss pathologies, or increased demand on iron storage as during normal pregnancy. Simple, direct and automation-ready procedures for measuring iron concentrations find wide applications in research, drug discovery and environmental monitoring. BioAssay Systems iron assay kit is designed to measure total iron directly in serum without any pretreatment. The improved method utilizes a chromogen that forms a blue colored complex specifically with Fe2+. Fe3+ in the sample is reduced to Fe2+, thus allowing the assay for total iron concentration. The intensity of the color, measured at 590nm, is directly proportional to the iron concentration in the sample.

1. I would like to know how serum samples should be prepared for use in the assay.



Sera are usually obtained by centrifugation of clotted blood (fresh blood standing for about 30 min at room temperature). Both, fresh serum or frozen serum can be assayed directly (50 µL sample + 200 µL working reagent).

2. Do DTT or IGEPAL/NP-40 in cell lysis buffer will have any influence on the assay?



Our data show that up to 4% NP-40 has no effect on the DIFE-250 assay. Up to 20 mM β-mercaptoethanol also had no effect on the DIFE-250. We did not test DTT, but we believe DTT would not interfere with this assay either.

3. Can you use DIFE-250 to measure iron in whole blood?



No, whole blood needs to be diluted to a degree that the normal serum iron concentration is below the detection limit of the assay. The assay is not suitable for measuring hemoglobin bound iron.

4. Please let us know the preparation for the liver tissue.



Tissue samples should be dried overnight at 106°C and weighed. Samples can then be solubilized in 6N nitric acid by heating at 100°C to release protein-associated iron. The solution is neutralized with NaOH, diluted in deionized water as necessary, and assayed for iron concentration using the DIFE-250 kits.

For more detailed product information and questions, please feel free to Contact Us. Or for more general information regarding our assays, please refer to our General Questions
Dichtl, S., Demetz, E., Haschka, D., Tymoszuk, P., Petzer, V., Nairz, M. & Theurl, I. (2019). Dopamine Is a Siderophore-Like Iron Chelator That Promotes Salmonella enterica Serovar Typhimurium Virulence in Mice. mBio, 10(1), e02624-18. Assay: Iron in mice serum.

Malhotra, H., Kumar, M., Chauhan, A. S., Dhiman, A., Chaudhary, S., Patidar, A. & Raje, M. (2019). Moonlighting Protein Glyceraldehyde-3-Phosphate Dehydrogenase: A Cellular Rapid-Response Molecule for Maintenance of Iron Homeostasis in Hypoxia. Cellular physiology and biochemistry: international journal of experimental cellular physiology, biochemistry, and pharmacology, 52(3), 517-531. Assay: Iron in mouse cells.

Cheli, V. T., Gonzalez, D. A. S., Marziali, L. N., Zamora, N. N., Guitart, M. E., Spreuer, V. & Paez, P. M. (2018). The Divalent Metal Transporter 1 (DMT1) is required for iron uptake and normal development of oligodendrocyte progenitor cells. Journal of Neuroscience, 38(43), 9142-9159. Assay: Iron in mice cells.

Efird, W. M., Fletcher, A. G., Draeger, R. W., Spang, J. T., Dahners, L. E., & Weinhold, P. S. (2018). Deferoxamine-Soaked Suture Improves Angiogenesis and Repair Potential After Acute Injury of the Chicken Achilles Tendon. Orthopaedic journal of sports medicine, 6(10), 2325967118802792. Assay: Iron in broiler chicks DFO.

Samba Mondonga, M., Calve, A., Mallette, F. A., & Santos, M. M. (2018). MyD88 regulates the expression of SMAD4 and the iron regulatory hormone hepcidin in hepatoma cells. Frontiers in cell and developmental biology, 6, 105. Assay: Iron in human kidney cells.

Worley, B. L., Kim, Y. S., Mardini, J., Zaman, R., Leon, K. E., Vallur, P. G. & Phaeton, R. (2018). GPx3 supports ovarian cancer progression by manipulating the extracellular redox environment. Redox Biology pii: S2213-2317(18)30891-7. Assay: Iron in human ovarian cells.

Asshoff, M., Petzer, V., Warr, M. R., Haschka, D., Tymoszuk, P., Demetz, E. & Fowles, P. (2017). Momelotinib inhibits ACVR1/ALK2, decreases hepcidin production, and ameliorates anemia of chronic disease in rodents. Blood, 129(13), 1823-1830. Assay: Iron in rat/mice serum.

Ma, X., Pham, V. T., Mori, H., MacDougald, O. A., Shah, Y. M., & Bodary, P. F. (2017). Iron elevation and adipose tissue remodeling in the epididymal depot of a mouse model of polygenic obesity. PloS one, 12(6), e0179889. Assay: Iron in mice serum.

Marks, E. S., Bonnemaison, M. L., Brusnahan, S. K., Zhang, W., Fan, W., Garrison, J. C., & Boesen, E. I. (2017). Renal iron accumulation occurs in lupus nephritis and iron chelation delays the onset of albuminuria. Scientific reports, 7(1), 12821. Assay: Iron in mice plasma.

Quan, Y. Y., Liu, Y. H., Lin, C. M., Wang, X. P., & Chen, T. S. (2017). Peroxynitrite dominates sodium nitroprusside-induced apoptosis in human hepatocellular carcinoma cells. Oncotarget 8(18): 29833-29845. Assay: Iron in cells.

Shang, Y. M., Wang, G. S., Sliney, D. H., Yang, C. H., & Lee, L. L. (2017). Light-emitting-diode induced retinal damage and its wavelength dependency in vivo. International journal of ophthalmology 10(2): 191-202. Assay: Iron in Sprague Dewley rats retina protein.

Sun, X., Zhao, Y., Jia, J., Xie, J., Cheng, J., Liu, H. & Fu, Y. (2017). Uninterrupted expression of CmSIT1 in a sclerotial parasite Coniothyrium minitans leads to reduced growth and enhanced antifungal ability. Frontiers in microbiology, 8, 2208. Assay: Iron in Coniothyrium minitans cells.

Szemraj, M., Oszajca, K., Szemraj, J., & Jurowski, P. (2017). MicroRNA expression analysis in serum of patients with congenital hemochromatosis and age-related macular degeneration (AMD). Medical science monitor: international medical journal of experimental and clinical research 23: 4050-4060. Assay: Iron in human serum.

Thongdee, P., & Na-Bangchang, K. (2017). The role of heme-oxygenase-1 in pathogenesis of cerebral malaria in the co-culture model of human brain microvascular endothelial cell and ITG Plasmodium falciparum-infected red blood cells. Asian Pacific journal of tropical medicine 10(1): 20-24. Assay: Iron in human brain cells.

Zhang, C. W., Tai, Y. K., Chai, B. H., Chew, K. C., Ang, E. T., Tsang, F. & Lim, K. L. (2017). Transgenic mice overexpressing the divalent metal transporter 1 exhibit iron accumulation and enhanced parkin expression in the brain. Neuromolecular medicine, 19(2-3), 375-386. Assay: Iron in mouse feces.

Hendricks, M. R., Lashua, L. P., Fischer, D. K., Flitter, B. A., Eichinger, K. M., Durbin, J. E. & Bomberger, J. M. (2016). Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity. Proceedings of the National Academy of Sciences, 113(6), 1642-1647. Assay: Iron in human cells.

Karoopongse, E., Marcondes, A. M., Yeung, C., Holman, Z., Kowdley, K. V., Campbell, J. S., & Deeg, H. J. (2016). Disruption of iron regulation after radiation and donor cell infusion. Biology of Blood and Marrow Transplantation, 22(7), 1173-1181. Assay: Iron in mouse serum.

Li, Y., Pan, K., Chen, L., Ning, J. L., Li, X., Yang, T. & Tao, G. (2016). Deferoxamine regulates neuroinflammation and iron homeostasis in a mouse model of postoperative cognitive dysfunction. Journal of neuroinflammation, 13(1), 268. Assay: Iron in mice cells.

Noguchi-Sasaki, M., Sasaki, Y., Shimonaka, Y., Mori, K., & Fujimoto-Ouchi, K. (2016). Treatment with anti-IL-6 receptor antibody prevented increase in serum hepcidin levels and improved anemia in mice inoculated with IL-6-producing lung carcinoma cells. BMC Cancer 16: 270. Assay: Iron in mouse serum.

Tamayo, E., Benabdellah, K., & Ferrol, N. (2016). Characterization of three new glutaredoxin genes in the arbuscular mycorrhizal fungus Rhizophagus irregularis: putative role of RiGRX4 and RiGRX5 in iron homeostasis. PloS one, 11(2), e0149606. Assay: Iron in yeast cells.

Theurl, I., Hilgendorf, I., Nairz, M., Tymoszuk, P., Haschka, D., Asshoff, M. & Sopper, S. (2016). On-demand erythrocyte disposal and iron recycling requires transient macrophages in the liver. Nature medicine, 22(8), 945. Assay: Iron in human plasma.

Anderson ER, et al (2011). Intestinal hypoxia-inducible factor-2alpha (HIF-2alpha) is critical for efficient erythropoiesis. J Biol Chem. 286(22):19533-40. Assay: Iron in mice serum.

Hamlin F, Latunde-Dada GO (2011). Iron bioavailibity from a tropical leafy vegetable in anaemic mice. Nutr Metab (Lond). 8:9. Assay: Iron in mice serum.

Waheed MM, et al (2011). Some Biochemical Characteristics and Preservation of Epididymal Camel Spermatozoa (Camelus dromedarius). Theriogenology 76(6):1126-33. Assay: Iron in camel epididymal fluid.

Ringseis R, et al (2010). Low availability of carnitine precursors as a possible reason for the diminished plasma carnitine concentrations in pregnant women. BMC Pregnancy Childbirth.10:17. Assay: Iron in human plasma.

Zhang Y, et al (2010). Diphthamide biosynthesis requires an organic radical generated by an iron-sulphur enzyme. Nature 465(7300):891-6. Assay: Iron in bacterial cell .

Zhu W, et al (2010). Genetic iron chelation protects against proteasome inhibition-induced dopamine neuron degeneration. Neurobiol Dis. 37(2):307-13. Assay: Iron in human neuron cell.

Chen H, et al (2009). Changes in iron-regulatory proteins in the aged rodent neural retina. Neurobiol Aging. 30(11):1865-76. Assay: Iron in rat serum.

Chen H, et al (2009). Dysfunction of the retinal pigment epithelium with age: increased iron decreases phagocytosis and lysosomal activity. Invest Ophthalmol Vis Sci. 50(4):1895-902. Assay: Iron in rat serum.

Shah, YM et al (2009). Intestinal hypoxia-inducible transcription factors are essential for iron absorption following iron deficiency. Cell Metab. 9(2):152-64. Assay: Iron in mouse erythrocytes.

Sudarshan S, et al (2009). Fumarate hydratase deficiency in renal cancer induces glycolytic addiction and hypoxia-inducible transcription factor 1alpha stabilization by glucose-dependent generation of reactive oxygen species. Mol Cell Biol. 29(15):4080-90. Assay: Iron in human tumor cell.

Yokosho K, et al (2009). OsFRDL1 is a citrate transporter required for efficient translocation of iron in rice. Plant Physiol. 149(1):297-305. Assay: Iron in plant rice.

Bandyopadhyay S, et al (2008). A proposed role for the Azotobacter vinelandii NfuA protein as an intermediate iron-sulfur cluster carrier. J Biol Chem. 283(20):14092-9. Assay: Iron in bacterial cell protein.

Raulfs EC, et al (2008). In vivo iron-sulfur cluster formation. Proc Natl Acad Sci U S A. 105(25):8591-6. Assay: Iron in bacterial cell protein.

Habel ME, Jung D. (2006) c-Myc over-expression in Ramos Burkitt's lymphoma cell line predisposes to iron homeostasis disruption in vitro. Biochem Biophys Res Commun. 341(4):1309-16. Assay: Iron in human lymphoma cell.

To find more recent publications, please click here.
Please inquire or request assay service or call 1-510-782-9988 x 2.
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QuantiChrom™ Iron Assay Kit
Catalog No: DIFE-250
Price: $329    Qty:
For orders of 10 or more kits, please call 1-510-7829988x1 or email us for best pricing and/or bulk order.

Shipping: RT
Shipment: Fedex Service
Delivery: 1-2 days (US), 3-6 days (Intl) Storage: 4°C

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