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Iron Assay Kit

BCA Serum, Soil
Catalog No. ABIN1000262
  • Target
    Iron
    Application
    Biochemical Assay (BCA)
    Sample Type
    Serum, Soil
    Specificity
    27 μg/dL (4.8 μM)
    Characteristics
    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. Cuvet or 96-well plate assay.
    Low interference in biological samples.
    No pretreatments are needed. Assays can be directly performed on serum samples.
    Components
    Reagent A: 50 mL. Reagent B: 4 mL. Reagent C: 4 mL. Iron Standard: 1 mL 10 mg/dL Fe2+.
    Material not included
    Pipeting devices and accessories. Clear bottom 96-well plates (e.g. Corning Costar) and plate reader. Cuvets and spectrophotometer for measuring OD at 510-630nm.
  • Application Notes
    Direct Assays: iron in biological samples (e.g. serum).
    Drug Discovery/Pharmacology: effects of drugs on iron metabolism. Environmental Monitoring: iron in soil extracts, mineralized samples.
    Protocol
    Procedure using 96-well plate:
    1. Standards. Prepare 400 µL 1000 µg/dL Premix by mixing 40 µL 10 mg/dL standard and 360 µL distilled water.
    2. Set up standards and samples. Transfer 50 µL diluted Standards and 50 µL samples into wells of a clear bottom 96-well plate. As a sample blank control, add 200 µL Reagent A to sample blank wells. Add 200 µL Working Reagent to all other wells. Tap plate to mix. Store diluted standards at 4 °C for future use.
    3. Incubate 40 min at room temperature and read optical density at 510- 630nm (peak absorbance at 590nm).

    Procedure using cuvette:
    1. Prepare standards as in 96-well assay. Set up centrifuge tubes labeled Standards and Samples (Sample and sample Blank). Transfer 250 µL Standards and Samples to tubes.
    2. Add 1000 µL Reagent A to Blank and 1000 µL Working Reagent to all other tubes. Mix by vortexing. Incubate 40 min at room temperature.
    3. Transfer to cuvets and read OD at 590nm (510nm-630nm).
    Reagent Preparation

    Reagent: Prepare enough Working Reagent by mixing 20 volumes of Reagent A, 1 volume Reagent B and 1 volume Reagent C. Fresh reconstitution is recommended. Equilibrate to room temperature before assay.

    Calculation of Results

    Subtract blank OD (water, #8) from the standard OD values and plot the OD against standard iron concentrations. Determine the slope using linear regression fitting. Typical serum iron values: 70-180 µg/dL.
    Conversions: 1 mg/dL Fe equals 179 µM, 0.001% or 10 ppm.

    Restrictions
    For Research Use only
  • Storage
    4 °C
  • Hamlin, Latunde-Dada: "Iron bioavailibity from a tropical leafy vegetable in anaemic mice." in: Nutrition & metabolism, Vol. 8, pp. 9, (2011) (PubMed).

    Anderson, Xue, Shah: "Intestinal hypoxia-inducible factor-2alpha (HIF-2alpha) is critical for efficient erythropoiesis." in: The Journal of biological chemistry, Vol. 286, Issue 22, pp. 19533-40, (2011) (PubMed).

    Waheed, Al-Eknah, El-Bahr: "Some biochemical characteristics and preservation of epididymal camel spermatozoa (Camelus dromedarius)." in: Theriogenology, Vol. 76, Issue 6, pp. 1126-33, (2011) (PubMed).

    Zhu, Li, Xie, Luo, Kaur, Andersen, Jankovic, Le: "Genetic iron chelation protects against proteasome inhibition-induced dopamine neuron degeneration." in: Neurobiology of disease, Vol. 37, Issue 2, pp. 307-13, (2010) (PubMed).

    Ringseis, Hanisch, Seliger, Eder: "Low availability of carnitine precursors as a possible reason for the diminished plasma carnitine concentrations in pregnant women." in: BMC pregnancy and childbirth, Vol. 10, pp. 17, (2010) (PubMed).

    Zhang, Zhu, Torelli, Lee, Dzikovski, Koralewski, Wang, Freed, Krebs, Ealick, Lin: "Diphthamide biosynthesis requires an organic radical generated by an iron-sulphur enzyme." in: Nature, Vol. 465, Issue 7300, pp. 891-6, (2010) (PubMed).

    Chen, Liu, Lukas, Suyeoka, Wu, Neufeld: "Changes in iron-regulatory proteins in the aged rodent neural retina." in: Neurobiology of aging, Vol. 30, Issue 11, pp. 1865-76, (2009) (PubMed).

    Yokosho, Yamaji, Ueno, Mitani, Ma: "OsFRDL1 is a citrate transporter required for efficient translocation of iron in rice." in: Plant physiology, Vol. 149, Issue 1, pp. 297-305, (2009) (PubMed).

    Shah, Matsubara, Ito, Yim, Gonzalez: "Intestinal hypoxia-inducible transcription factors are essential for iron absorption following iron deficiency." in: Cell metabolism, Vol. 9, Issue 2, pp. 152-64, (2009) (PubMed).

    Chen, Lukas, Du, Suyeoka, Neufeld: "Dysfunction of the retinal pigment epithelium with age: increased iron decreases phagocytosis and lysosomal activity." in: Investigative ophthalmology & visual science, Vol. 50, Issue 4, pp. 1895-902, (2009) (PubMed).

    Sudarshan, Sourbier, Kong, Block, Valera Romero, Yang, Galindo, Mollapour, Scroggins, Goode, Lee, Gourlay, Trepel, Linehan, Neckers: "Fumarate hydratase deficiency in renal cancer induces glycolytic addiction and hypoxia-inducible transcription factor 1alpha stabilization by glucose-dependent generation of reactive oxygen species." in: Molecular and cellular biology, Vol. 29, Issue 15, pp. 4080-90, (2009) (PubMed).

    Bandyopadhyay, Naik, OCarroll, Huynh, Dean, Johnson, Dos Santos: "A proposed role for the Azotobacter vinelandii NfuA protein as an intermediate iron-sulfur cluster carrier." in: The Journal of biological chemistry, Vol. 283, Issue 20, pp. 14092-9, (2008) (PubMed).

    Raulfs, OCarroll, Dos Santos, Unciuleac, Dean: "In vivo iron-sulfur cluster formation." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 105, Issue 25, pp. 8591-6, (2008) (PubMed).

    Habel, Jung: "c-Myc over-expression in Ramos Burkitt's lymphoma cell line predisposes to iron homeostasis disruption in vitro." in: Biochemical and biophysical research communications, Vol. 341, Issue 4, pp. 1309-16, (2006) (PubMed).

  • Target
    Iron
    Target Type
    Element
    Background
    Quantitative determination of iron ions Fe3+ and/or Fe2+ by colorimetric (590nm) method.
    Procedure: 30 min.

    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. This 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. The optimized formulation substantially reduces interference by substances in the raw samples.
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