What laboratory tests should be ordered to rule out (exclude) anemia?

Laboratory Tests to Rule Out Anemia

Start with a complete blood count (CBC) with hemoglobin and hematocrit to diagnose anemia, then immediately add reticulocyte count, iron studies (serum ferritin, transferrin saturation, serum iron), vitamin B12, folate, and C-reactive protein (CRP) to determine the underlying cause. 1, 2

Initial Screening Tests

Complete Blood Count (CBC) with Indices

• Hemoglobin and hematocrit are the primary tests to confirm anemia, with cutoffs of <13 g/dL in men and <12 g/dL in women (or <11 g/dL in pregnant women). 3, 2
• Mean corpuscular volume (MCV) classifies anemia as microcytic (<80 fL), normocytic (80-100 fL), or macrocytic (>100 fL), which narrows the differential diagnosis significantly. 3, 2
• Red cell distribution width (RDW) helps distinguish iron deficiency (elevated RDW) from thalassemia (normal RDW) in microcytic anemia. 2, 4
• Mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC) provide additional clues, with low values suggesting iron deficiency or thalassemia. 2, 5

Reticulocyte Count

• Reticulocyte count is essential and should be ordered immediately upon detecting anemia to distinguish production defects (low/normal reticulocytes) from hemolysis or blood loss (elevated reticulocytes). 1, 2, 4
• A low or inappropriately normal reticulocyte count in an anemic patient indicates bone marrow failure to respond, suggesting iron deficiency, vitamin B12/folate deficiency, anemia of chronic disease, or primary bone marrow disease. 1, 6
• An elevated reticulocyte count excludes nutritional deficiencies and suggests active bleeding, hemolysis, or hemoglobinopathies. 1, 2

Iron Status Assessment

Core Iron Studies

• Serum ferritin is the most sensitive marker for iron stores, with <30 μg/L indicating absolute iron deficiency in non-inflammatory states. 3, 2
• Transferrin saturation (TSAT) <20% indicates functional or absolute iron deficiency and represents iron available for erythropoiesis. 3, 2
• Serum iron and total iron-binding capacity (TIBC) help calculate transferrin saturation and distinguish iron deficiency (low iron, high TIBC) from anemia of chronic disease (low iron, low TIBC). 3, 4

Critical Interpretation Pitfall

• Ferritin is an acute phase reactant and can be falsely elevated in inflammation, infection, or malignancy. 3, 2
• In inflammatory states, ferritin 30-100 μg/L may still represent iron deficiency; consider ferritin <100 μg/L with TSAT <20% as iron deficient in these patients. 3, 2
• Always measure CRP alongside iron studies to assess for inflammation that may confound ferritin interpretation. 3, 2

Nutritional Deficiency Markers

Vitamin B12 and Folate

• Vitamin B12 and folate levels must be checked before initiating treatment, as deficiency causes macrocytic anemia and can coexist with iron deficiency. 3, 6, 7
• Vitamin B12 deficiency allowed to progress >3 months produces permanent spinal cord degeneration that folic acid supplementation will not prevent. 6
• Hematocrit and reticulocyte counts should be monitored during treatment; if reticulocytes don't increase or remain elevated, repeat iron and folate studies to identify complicating deficiencies. 6

Inflammatory Markers

C-Reactive Protein (CRP)

• CRP is mandatory to identify anemia of chronic disease and interpret ferritin levels correctly in inflammatory conditions. 3, 2
• Elevated CRP with elevated ferritin (>100 μg/L) but low TSAT suggests anemia of chronic disease or functional iron deficiency from hepcidin upregulation. 3, 7

Additional Tests Based on Clinical Context

Renal Function

• Serum creatinine and estimated glomerular filtration rate (eGFR) should be checked, as chronic kidney disease causes anemia through decreased erythropoietin production. 3
• In CKD patients with anemia, screen at minimum yearly with hemoglobin measurement. 3

Thyroid Function

• Thyroid function tests are part of routine assessment in patients with anemia, particularly those with heart failure or unexplained normocytic anemia. 3

Hemoglobin Electrophoresis

• Order when thalassemia is suspected (microcytic anemia with normal or elevated ferritin, family history, or specific ethnic background). 2, 4

Algorithmic Approach

1. Order CBC with indices → Confirms anemia and provides MCV classification
2. Add reticulocyte count immediately → Distinguishes production vs. destruction/loss
3. Order iron panel (ferritin, TSAT, serum iron) + CRP → Identifies iron deficiency and inflammation
4. Add vitamin B12 and folate → Rules out nutritional deficiencies
5. Include creatinine/eGFR → Assesses for CKD contribution
6. Consider thyroid function tests → Especially in unexplained cases

Common Pitfalls to Avoid

• Never rely on hemoglobin/hematocrit alone without checking iron studies, as normal CBC parameters can coexist with iron depletion. 8
• Don't interpret ferritin in isolation—always consider inflammatory status via CRP, as ferritin >100 μg/L doesn't exclude iron deficiency in inflammation. 3, 2
• Don't assume a "normal" reticulocyte count is adequate—it may be inappropriately low for the degree of anemia, indicating inadequate bone marrow response. 1
• Never give folic acid without checking B12 first, as folic acid can mask B12 deficiency anemia while allowing irreversible neurologic damage to progress. 6
• Don't miss coexisting deficiencies—high RDW suggests mixed deficiency states even when MCV is normal. 2