Enzyme Immunoassay for the Quantitative Determination of Ferritin Concentration in Human Serum
Introduction
Ferritin is the primary iron storage protein in the human body, with a molecular weight of approximately 450 kilo Daltons (kDa). It is found in various tissues, including the liver, spleen, brain, and bone marrow. The ferritin molecule comprises 24 protein subunits that form a central hollow sphere capable of storing up to 4,000 iron atoms in a soluble form. Approximately 1% of total plasma iron is bound to ferritin, making it the most important and highly specific iron storage form in cells (Beard, 2001). When iron deficiency occurs, ferritin-bound iron is rapidly mobilized to maintain iron homeostasis.
Ferritin levels are high at birth, gradually decrease in infancy, and remain low during childhood. During puberty, ferritin levels increase steadily, and in adulthood, men generally exhibit higher ferritin concentrations than women (Cook et al., 1974). Serum ferritin is a reliable biomarker for assessing iron storage levels as it remains relatively stable compared to fluctuating iron values.
Clinical Significance
Ferritin level determination is a critical parameter in diagnosing and monitoring iron-related disorders. Low ferritin levels are indicative of iron deficiency anemia, whereas elevated ferritin levels are associated with conditions such as hemochromatosis, chronic disease anemia, hemolytic anemias (e.g., thalassemia), liver disease, malignancies, and inflammation (Powell et al., 1998). Regular ferritin level assessments are recommended for at-risk populations, including blood donors, pregnant women, hemodialysis patients, and infants (Dawson et al., 1992).
Test Principle
The Ferritin Quantitative Test Kit is based on a solid-phase enzyme-linked immunosorbent assay (ELISA). This assay utilizes a monoclonal anti-ferritin antibody immobilized onto microtiter wells and a horseradish peroxidase (HRP)-conjugated monoclonal anti-ferritin antibody as the detection reagent. The test sample reacts with the immobilized antibodies. After incubation and washing, the enzyme conjugate is added, forming a sandwich complex between the solid-phase and conjugated antibodies. Following another wash step, a chromogen-substrate solution is added, leading to a blue color development. The reaction is stopped using a stop solution, turning the color yellow. The absorbance at 450 nm is directly proportional to the ferritin concentration in the sample.
References
- Beard, J. L. (2001). Iron biology in immune function, muscle metabolism, and neuronal functioning. Journal of Nutrition, 131(2), 565S-580S.
- Cook, J. D., Lipschitz, D. A., Miles, L. E. M., & Finch, C. A. (1974). Serum ferritin as a measure of iron stores in normal subjects. American Journal of Clinical Nutrition, 27, 681-687.
- Dawson, D. W., et al. (1992). The accuracy and chemical interpretation of serum ferritin assay. Clinical Laboratory Haematology, 14(1), 47-52.
- Powell, L. W., et al. (1998). Diagnosis of hemochromatosis. Annals of Internal Medicine, 129, 925-931.