What laboratory tests should be ordered to evaluate anemia?

Laboratory Evaluation of Anemia

Order a complete blood count with red cell indices (MCV, MCH, RDW), absolute reticulocyte count, serum ferritin, transferrin saturation, and vitamin B12 and folate levels as your initial anemia workup. 1, 2

Essential First-Line Tests

Complete Blood Count with Indices

• Hemoglobin and hematocrit define anemia severity (hemoglobin < 13 g/dL in men, < 12 g/dL in non-pregnant women), with hemoglobin preferred for its superior reproducibility across laboratories. 1
• Mean corpuscular volume (MCV) classifies anemia as microcytic (< 80 fL), normocytic (80-100 fL), or macrocytic (> 100 fL), guiding your differential diagnosis. 2
• Mean corpuscular hemoglobin (MCH) is more reliable than MCHC for detecting iron deficiency because it is less dependent on storage conditions and laboratory equipment. 3
• Red cell distribution width (RDW) is critical for detecting mixed deficiencies, with RDW > 14% suggesting iron deficiency or combined nutritional deficits even when MCV appears normal. 2
• White blood cell count with differential and platelet count assess bone marrow function, with abnormalities in two or more cell lines warranting hematology consultation. 4, 1

Reticulocyte Assessment

• Absolute reticulocyte count distinguishes impaired erythropoiesis (low/normal count) from hemolysis or blood loss (elevated count), making it essential for determining the pathophysiologic mechanism. 1, 2
• A low reticulocyte count in the setting of anemia indicates inadequate bone marrow response, most commonly from iron deficiency, nutritional deficiencies, or insufficient erythropoietin production. 4

Iron Status Evaluation

• Serum ferritin is the single most powerful test for iron stores, with levels < 30 μg/L confirming iron deficiency in the absence of inflammation. 1, 2
• Transferrin saturation (TSAT) < 15-20% supports iron deficiency and is less affected by inflammatory states than ferritin, making it more reliable in patients with chronic disease. 3, 1
• C-reactive protein (CRP) should be measured alongside ferritin because ferritin rises as an acute-phase reactant during inflammation, potentially masking true iron deficiency. 3, 2
• In inflammatory conditions, use a higher ferritin threshold (< 100 μg/L) to diagnose iron deficiency. 2

Vitamin Deficiency Testing

• Vitamin B12 and folate levels are essential components of the initial workup, particularly for evaluating macrocytic anemia. 1, 2
• These tests are not significantly affected by inflammatory conditions, unlike iron studies. 2

Algorithmic Approach Based on Initial Results

If MCV < 80 fL (Microcytic)

• Iron deficiency is the most common cause globally. 1
• Confirm with ferritin < 30 μg/L and TSAT < 20%. 3, 1
• In adult men and postmenopausal women with confirmed iron deficiency, investigate for gastrointestinal bleeding. 3, 1
• If iron studies are normal, obtain hemoglobin electrophoresis to evaluate for thalassemia trait, particularly in patients of Mediterranean, African, or Southeast Asian descent. 3

If MCV 80-100 fL (Normocytic)

• Evaluate reticulocyte count first. 1
• If reticulocyte count is elevated, assess for hemolysis with haptoglobin, lactate dehydrogenase (LDH), and indirect bilirubin. 3, 1
• If reticulocyte count is low/normal, consider anemia of chronic inflammation (elevated ferritin with low TSAT) or early nutritional deficiencies. 3, 1

If MCV > 100 fL (Macrocytic)

• Measure vitamin B12 and folate levels immediately. 3, 1
• Obtain peripheral blood smear to look for hypersegmented neutrophils (megaloblastic pattern) or oval macrocytes. 3
• Review medications, focusing on thiopurines, chemotherapy agents, anticonvulsants, and methotrexate. 3
• If reticulocyte count is elevated, evaluate for hemolysis. 3

Additional Tests in Specific Contexts

When Initial Workup is Unrevealing

• Peripheral blood smear provides morphologic clues that automated indices may miss. 3, 1
• Bone marrow aspiration and biopsy should be considered when pancytopenia is present or when the initial workup fails to establish a diagnosis. 1

Specialized Iron Tests (Not Routinely Available)

• Reticulocyte hemoglobin content (CHr) and percent hypochromic red blood cells (PHRBC) have sensitivities and specificities equal to or greater than ferritin and TSAT for identifying functional iron deficiency, but require specialized equipment. 4, 2
• These tests are particularly useful in chronic kidney disease patients on dialysis. 4

Critical Pitfalls to Avoid

• Do not rely solely on MCV for classification, as coexisting iron deficiency and vitamin B12 deficiency can neutralize MCV changes, yielding a falsely normal value with elevated RDW. 3, 2
• Do not misinterpret ferritin without assessing inflammatory status, as ferritin can be falsely elevated in inflammation, chronic disease, malignancy, or liver disease. 3, 1, 2
• Do not overlook mixed deficiencies, which are common and require both iron studies and vitamin levels to be checked simultaneously. 3
• Do not fail to investigate the underlying cause of confirmed deficiencies, particularly gastrointestinal bleeding in iron deficiency or malabsorption syndromes in vitamin deficiencies. 3, 1

Timing Considerations for Specific Populations

• In hemodialysis patients, measure hemoglobin levels before dialysis (not after) because postdialysis hemoglobin varies with ultrafiltration volume, increasing approximately 3-4 g/dL for every liter removed. 4
• Preferably obtain levels before the midweek dialysis session, as hemoglobin levels vary throughout the week. 4