Overview
For quantitative determination of iron ions Fe 3+ and/or Fe 2+ and evaluation of drug effects on iron metabolism. The assay uses OD590nm for signal readout. Compatible sample input includes Biological (e.g. serum) and environmental samples. Typical stated assay timing is 30 min.
Key elements and design rationale
- Readout format: OD590nm supports plate-based signal acquisition and consistent comparison across matched samples.
- Sample compatibility: The stated sample scope includes Biological (e.g. serum) and environmental samples, which is useful when aligning matrix type with calibration and control design.
- Analytical range context: The supplied specifications include a stated detection limit of 27 µg/dL (4.8 µM) for interpreting low-signal samples.
- Feature emphasis: Sensitive and accurate. Linear detection range 27 µg/dL (4.8 µM) to 1,000 µg/dL (179 µM) iron in a 96-well plate assay.
Additional feature notes highlight Simple and high-throughput. The procedure involves the 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 the reagent and signal stability. Cuvette or 96-well plate assay. Available format information for this listing includes 250 Tests.
Biological background
This product is centered on measurement of iron within the matrices described for the assay. In practice, datasets from this type of format are typically interpreted by comparing relative signal, activity, or abundance across matched control and experimental groups rather than relying on a single value in isolation. Careful alignment of sample matrix, incubation window, and calibration strategy is important when comparing results across plates, operators, or study days.
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 for iron storage 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.
Detection method
Colorimetric (OD 590 nm).
Detection limit and analytical sensitivity
Reported detection limit: 27 µg/dL (4.8 µM).
Procedures and timing
Stated procedure or timing information: 30 min.
Research relevance and current trends
- Plate-based quantification and side-by-side group comparison remain central use cases for this assay format.
- The product notes emphasize multi-sample throughput, making it relevant for screening-oriented and larger batch comparison studies.
- The description supports intervention-focused study designs in which researchers compare baseline and perturbed conditions.
Common research applications
- Quantify iron in biological (serum) by OD590 nm readout.
- Compare treatment or phenotype groups using matched biological (serum) handling.
- Monitor time-course or pre/post changes in biological (serum) across study conditions.
Interpretation is usually strongest when signal changes are assessed alongside matrix-matched controls, replicate agreement, and the assay's stated analytical window.
Notes for experimental interpretation
- Matrix composition, background signal, and sample handling can influence apparent response; compare like-with-like whenever possible.
- Use appropriate blanks, controls, and replicate wells to distinguish biological differences from plate, reagent, or handling variability.
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).
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.
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.
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 laboratories requiring additional technical capacity, we provide scientific support services including assay execution, method guidance, product sourcing, and customization to align the assay with specific experimental objectives. If you need assistance selecting the appropriate kit configuration, adapting the workflow to your application, or identifying related research services, please click Talk to a Scientist, email support@biohippo.com, or review our Research Services; a member of our scientific team will follow up with recommendations tailored to your study.
Extracellular vesicles promote transkingdom nutrient transfer during viral-bacterial co-infection
Hendricks, MR et al (2021). Extracellular vesicles promote transkingdom nutrient transfer during viral-bacterial co-infection. Cell Reports, 34(4), 108672. Assay: Iron in human extracelllular vesicles.
The neurophysiological effects of iron in early life stages of zebrafish
Hassan AT, Kwong R (2020). The neurophysiological effects of iron in early life stages of zebrafish. Environmental Pollution, 267, 115625. Assay: Iron in water.
Identification of pannexin 2 as a novel marker correlating with ferroptosis and malignant phenotypes of prostate cancer cells
Liao, D et al (2020). Identification of pannexin 2 as a novel marker correlating with ferroptosis and malignant phenotypes of prostate cancer cells. OncoTargets and Therapy, 13, 4411-4421. Assay: Iron in human prostate cancer cell lysates.
Modulating NF-kB, MAPK, and PI3K/AKT signaling by ergothioneine attenuates iron overload-induced hepatocellular injury in rats
Salama SA, Omar HA (2021). Modulating NF-kB, MAPK, and PI3K/AKT signaling by ergothioneine attenuates iron overload-induced hepatocellular injury in rats. Journal of Biochemical and Molecular Toxicology, e22729. Assay: Iron in rat.
Heme-dependent er stress apoptosis: A mechanism for the selective toxicity of the dihydroartemisinin, nsc735847, in colorectal cancer cells
Elhassanny, AEM et al (2020). Heme-dependent er stress apoptosis: A mechanism for the selective toxicity of the dihydroartemisinin, nsc735847, in colorectal cancer cells. Frontiers in Oncology, 10. Assay: Iron in human colon cancer cell lysates.
Ubiquitin-like modifier activating enzyme 1 as a novel diagnostic and prognostic indicator that correlates with ferroptosis and the malignant phenotypes of liver cancer cells
Shan, Y et al (2020). Ubiquitin-like modifier activating enzyme 1 as a novel diagnostic and prognostic indicator that correlates with ferroptosis and the malignant phenotypes of liver cancer cells. Frontiers in Oncology, 10, 592413. Assay: Iron in human liver cancer cell lysates.
Transcription analysis of the stress and immune response genes to temperature stress in ostrinia furnacalis
Chen, K et al (2019). Transcription analysis of the stress and immune response genes to temperature stress in ostrinia furnacalis. Frontiers in Physiology, 10, 1289. Assay: Iron in ostrinia furnacalis plasma.
Expression analysis of glutathione S-transferases and ferritins during the embryogenesis of the tick Haemaphysalis longicornis
Hernandez, EP et al. (2020). Expression analysis of glutathione S-transferases and ferritins during the embryogenesis of the tick Haemaphysalis longicornis. Heliyon, 6(3), e03644. Assay: Iron in haemaphysalis longicornis egg homogenates.
Salmonella effector SpvB aggravates dysregulation of systemic iron metabolism via modulating the hepcidin-ferroportin axis
Deng, Q et al (2021). Salmonella effector SpvB aggravates dysregulation of systemic iron metabolism via modulating the hepcidin-ferroportin axis. Gut Microbes, 13(1), 1-18. Assay: Iron in mouse serum and liver.
Ubiquitin-specific protease 7 promotes ferroptosis via activation of the p53/TfR1 pathway in the rat hearts after ischemia/reperfusion
Tang, LJ et al (2021). Ubiquitin-specific protease 7 promotes ferroptosis via activation of the p53/TfR1 pathway in the rat hearts after ischemia/reperfusion. Free Radical Biology & Medicine, 162, 339-352. Assay: Iron in rat cardiac tissue homogenates.
Iron homeostasis in a mouse model of thalassemia intermedia is altered between adolescence and adulthood
Sanyear, C et al (2020). Iron homeostasis in a mouse model of thalassemia intermedia is altered between adolescence and adulthood. PeerJ, 8, e8802. Assay: Iron in mouse serum.
Deficiency of Meis1, a transcriptional regulator, in mice and worms: Neurochemical and behavioral characterizations with implications in the restless legs syndrome
Lyu, S et al (2020). Deficiency of Meis1, a transcriptional regulator, in mice and worms: Neurochemical and behavioral characterizations with implications in the restless legs syndrome. Journal of Neurochemistry, 155(5), 522-537. Assay: Iron in mouse serum.
Cardiac-specific loss of mitoNEET expression is linked with age-related heart failure
Furihata, T et al (2021). Cardiac-specific loss of mitoNEET expression is linked with age-related heart failure. Communications Biology, 4(1), 138. Assay: Iron in mouse mitochondria.
Mu opioid receptor knockout mouse: Phenotypes with implications on restless legs syndrome
Lyu, S et al (2020). Mu opioid receptor knockout mouse: Phenotypes with implications on restless legs syndrome. Journal of Neuroscience Research, 98(8), 1532-1548. Assay: Iron in mouse.
Matte JJ, Audet I (2020) Maternal perinatal transfer of vitamins and trace elements to piglets. Animal, 14(1), 31-38. As
Matte JJ, Audet I (2020) Maternal perinatal transfer of vitamins and trace elements to piglets. Animal, 14(1), 31-38. Assay: Iron in pig serum.
Ferrous-iron-activated transcriptional factor adhr regulates redox homeostasis in clostridium beijerinckii
Yang, B et al (2020). Ferrous-iron-activated transcriptional factor adhr regulates redox homeostasis in clostridium beijerinckii. Applied and Environmental Microbiology, 86(7). Assay: Iron in clostridium beijerinckii AdHr protein.
Dek expression in breast cancer cells leads to the alternative activation of tumor associated macrophages
Pease, NA et al (2020). Dek expression in breast cancer cells leads to the alternative activation of tumor associated macrophages. Cancers, 12(7). Assay: Iron in mouse cell lysates.
The novel ECF56 SigG1-RsfG system modulates morphological differentiation and metal-ion homeostasis in Streptomyces tsukubaensis
Oliveira, R et al (2020). The novel ECF56 SigG1-RsfG system modulates morphological differentiation and metal-ion homeostasis in Streptomyces tsukubaensis. Scientific Reports, 10(1), 21728. Assay: Iron in streptomyces tsukubaensis cell culture supernatant.
Iron metabolism in the peripheral nervous system: The role of dmt1, ferritin, and transferrin receptor in schwann cell maturation and myelination
Santiago Gonzalez, DA et al (2019). Iron metabolism in the peripheral nervous system: The role of dmt1, ferritin, and transferrin receptor in schwann cell maturation and myelination. The Journal of Neuroscience: The Official Journal of the Society for Neuroscience, 39(50), 9940-9953. Assay: Iron in mouse DRG explant cultures and sciatic nerve homogenates.
Dopamine Is a Siderophore-Like Iron Chelator That Promotes Salmonella enterica Serovar Typhimurium Virulence in Mice
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.
Moonlighting Protein Glyceraldehyde-3-Phosphate Dehydrogenase: A Cellular Rapid-Response Molecule for Maintenance of Iron Homeostasis in Hypoxia
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.
The Divalent Metal Transporter 1 (DMT1) is required for iron uptake and normal development of oligodendrocyte progenitor 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.
MyD88 regulates the expression of SMAD4 and the iron regulatory hormone hepcidin in hepatoma cells
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.
GPx3 supports ovarian cancer progression by manipulating the extracellular redox environment
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.
Momelotinib inhibits ACVR1/ALK2, decreases hepcidin production, and ameliorates anemia of chronic disease in rodents
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.
Iron elevation and adipose tissue remodeling in the epididymal depot of a mouse model of polygenic obesity
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.
Renal iron accumulation occurs in lupus nephritis and iron chelation delays the onset of albuminuria
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.
Peroxynitrite dominates sodium nitroprusside-induced apoptosis in human hepatocellular carcinoma cells
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.
Light-emitting-diode induced retinal damage and its wavelength dependency in vivo
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.
Uninterrupted expression of CmSIT1 in a sclerotial parasite Coniothyrium minitans leads to reduced growth and enhanced antifungal ability
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.
Transgenic mice overexpressing the divalent metal transporter 1 exhibit iron accumulation and enhanced parkin expression in the brain
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.
Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity
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.
Disruption of iron regulation after radiation and donor cell infusion
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.
Deferoxamine regulates neuroinflammation and iron homeostasis in a mouse model of postoperative cognitive dysfunction
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.
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
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.
Characterization of three new glutaredoxin genes in the arbuscular mycorrhizal fungus Rhizophagus irregularis: putative role of RiGRX4 and RiGRX5 in iron homeostasis
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.
On-demand erythrocyte disposal and iron recycling requires transient macrophages in the liver
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.
Intestinal hypoxia-inducible factor-2alpha (HIF-2alpha) is critical for efficient erythropoiesis
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.
Iron bioavailibity from a tropical leafy vegetable in anaemic mice
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.
Low availability of carnitine precursors as a possible reason for the diminished plasma carnitine concentrations in pregnant women
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.
Diphthamide biosynthesis requires an organic radical generated by an iron-sulphur enzyme
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 .
Genetic iron chelation protects against proteasome inhibition-induced dopamine neuron degeneration
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.
Changes in iron-regulatory proteins in the aged rodent neural retina
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.
Dysfunction of the retinal pigment epithelium with age: increased iron decreases phagocytosis and lysosomal activity
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.
Intestinal hypoxia-inducible transcription factors are essential for iron absorption following iron deficiency
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.
Fumarate hydratase deficiency in renal cancer induces glycolytic addiction and hypoxia-inducible transcription factor 1alpha stabilization by glucose-dependent generation of reactive oxygen species
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.
OsFRDL1 is a citrate transporter required for efficient translocation of iron in rice
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.
A proposed role for the Azotobacter vinelandii NfuA protein as an intermediate iron-sulfur cluster carrier
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.
In vivo iron-sulfur cluster formation
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 dis
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.
Research budgets are tight — we get it. That's why we've put together a fresh round of exclusive promotions designed to help you stock up on the reagents, kits, and consumables your lab depends on, without stretching your budget.
🔬 What's on offer right now:
10% Off Pre-Designed siRNA Sets — use code SIRNA10
20% Off Transmembrane Proteins — use code TM20
$50 Off All ELISA Kits — auto-applied at checkout (code ELISA50)
50% Off Lab Consumables + Free Shipping on orders $500+
15% Off Proteins from Trusted Suppliers — use code PROTEIN15
$99 Pipette Filler Promotion Package (reg. $236)
Save 10% on your first order of any R&D grade DENARASE® with code DENARASE10 at checkout.
