Iron deficiency anemia (IDA) is the most prevalent anemia globally, affecting approximately 2 billion people. It results from insufficient iron to support normal erythropoiesis, leading to reduced hemoglobin synthesis and microcytic hypochromic RBCs. Understanding the laboratory diagnosis of IDA and distinguishing it from other microcytic anemias is a core skill in the hematology laboratory.
Iron Metabolism
Total body iron content is 3–5 g in adults. Approximately 65% is contained in hemoglobin (erythron iron), 10% in myoglobin and enzymes, 20–30% in storage (ferritin, hemosiderin), and less than 1% in transport (transferrin-bound). Dietary iron (1–2 mg/day) is absorbed in the duodenum by enterocytes via DMT1 (divalent metal transporter 1). Heme iron from animal sources has higher bioavailability (25–30%) than non-heme iron (5–10%). Iron absorption is regulated by hepcidin, a liver-derived peptide that degrades ferroportin (the sole iron export channel from enterocytes, macrophages, and hepatocytes). Hepcidin is increased by inflammation (IL-6) and iron overload, and decreased by iron deficiency, hypoxia, and erythropoietic drive (erythroferrone).
Stages of Iron Deficiency
Iron deficiency develops in three stages. Stage 1 (iron depletion): storage iron is exhausted — ferritin decreases but hemoglobin remains normal. Stage 2 (iron-deficient erythropoiesis): transport iron is insufficient — transferrin saturation falls below 16%, soluble transferrin receptor increases, and free erythrocyte protoporphyrin (FEP) rises. Hemoglobin begins to decrease. Stage 3 (iron deficiency anemia): hemoglobin falls below the reference range, MCV and MCH decrease (microcytic hypochromic), and the peripheral blood smear shows hypochromic microcytes with anisocytosis (elevated RDW).
Laboratory Diagnosis
Serum ferritin is the single most useful test: ferritin < 15–30 ng/mL is diagnostic of iron deficiency. Ferritin below 45 ng/mL has high sensitivity; however, ferritin is an acute-phase reactant and can be falsely normal or elevated in inflammation, infection, liver disease, and malignancy, reducing its specificity. Transferrin saturation (TSAT) = serum iron / TIBC × 100; TSAT < 16% suggests iron-deficient erythropoiesis. Total iron-binding capacity (TIBC) is elevated in iron deficiency (reflecting increased transferrin production). Soluble transferrin receptor (sTfR) is elevated in iron deficiency and unaffected by inflammation; the sTfR/log ferritin index helps distinguish IDA from anemia of chronic disease. CRP is measured concurrently to assess inflammation.
Peripheral Blood and CBC Findings
The CBC shows low hemoglobin (often < 10 g/dL in advanced IDA), low MCV (< 80 fL), low MCH (< 27 pg), low MCHC (< 33 g/dL), and elevated RDW (> 15%). The RDW is typically higher in IDA than in thalassemia trait — one of the key distinguishing features. The reticulocyte count is low for the degree of anemia (RPI < 2), reflecting inadequate iron for erythropoiesis. The reticulocyte hemoglobin content (RET-He) decreases early in iron-deficient erythropoiesis. The peripheral blood smear shows hypochromic (pale) microcytes, pencil-shaped cells (elliptocytes), and occasional target cells. In severe cases, bizarre poikilocytes and fragmented cells may be seen.
Causes of Iron Deficiency
In adults, chronic blood loss is the most common cause. Gastrointestinal bleeding (peptic ulcer, colon cancer, angiodysplasia, hemorrhoids, inflammatory bowel disease, hookworm infection) and menorrhagia (heavy menstrual bleeding) are the predominant sources. Decreased dietary intake or absorption occurs in vegans, gastric bypass surgery, Helicobacter pylori infection, and autoimmune gastritis. Increased iron requirements occur in pregnancy and lactation (total requirement ~1000 mg), infancy and childhood, and adolescence. In older adults, gastrointestinal malignancy must always be excluded.
Differential Diagnosis
Thalassemia trait (α or β) presents with microcytic hypochromic indices but normal or elevated RBC count, normal RDW, and normal iron studies (ferritin, TSAT). Hemoglobin electrophoresis shows elevated HbA2 in β-thalassemia trait. Anemia of chronic disease (ACD) is normocytic to mildly microcytic, with low TSAT but normal or high ferritin (inflammatory pattern), low sTfR, and elevated CRP. Sideroblastic anemia (congenital or acquired/MDS) shows a dimorphic RBC population on smear, elevated serum iron and ferritin, and ring sideroblasts on bone marrow iron stain. Lead poisoning causes microcytic anemia with basophilic stippling on smear and elevated blood lead levels.
Treatment and Response Monitoring
Oral iron therapy (ferrous sulfate 325 mg = 65 mg elemental iron, 1–3 times daily) is first-line treatment. Response is monitored by the reticulocyte count, which increases within 5–10 days (reticulocytosis, RPI > 2). Hemoglobin rises 1–2 g/dL over 2–4 weeks, and normalization typically occurs within 6–8 weeks. Iron therapy should continue for 3–6 months after Hb normalization to replete iron stores (ferritin > 50 ng/mL). Non-response is investigated for ongoing blood loss, malabsorption, non-adherence, or alternative diagnosis. Intravenous iron (iron sucrose, ferric carboxymaltose) is indicated for intolerance to oral iron, severe deficiency, malabsorption, or chronic kidney disease with ESA therapy.
