When should hemoglobin (Hb) electrophoresis be ordered?

When to Order Hemoglobin Electrophoresis

Order hemoglobin electrophoresis when you identify microcytosis with hypochromia in patients with normal iron studies, particularly in those from ethnic backgrounds at risk for hemoglobinopathies, to prevent unnecessary gastrointestinal investigations. 1

Primary Indications

Microcytic Anemia with Normal Iron Studies

• Order hemoglobin electrophoresis specifically when MCV is reduced with normal ferritin, transferrin saturation, and iron levels 1
• This prevents unnecessary GI workup in patients whose microcytosis stems from thalassemia rather than iron deficiency 1
• Pay particular attention when MCV is disproportionately low relative to the degree of anemia, which suggests thalassemia trait 1

Suspected Hemoglobinopathies

• Order when clinical presentation suggests sickle cell disease: normocytic normochromic anemia with hemolysis markers, painful crises, or family history 2
• Request comprehensive hemoglobinopathy screening using HPLC, capillary electrophoresis, mass spectrometry, or gel electrophoresis, confirmed by a second test 2, 3
• Never rely on sickle solubility testing alone - it cannot differentiate between heterozygous, compound heterozygous, or homozygous states 2

Unexplained Cyanosis or Methemoglobinemia

• Order when patients present with familial cyanosis or blue-tinged skin without cardiopulmonary disease 4
• Hemoglobin electrophoresis at pH 7.1 specifically identifies hemoglobin M variants caused by α-, β-, or γ-globin gene mutations 4
• However, recognize that DNA sequencing of α- and β-globin genes is technically simpler, more widely available, and more specific than electrophoresis for HbM variants 4

Technical Approach and Confirmatory Testing

Initial Screening Strategy

• Use HPLC or capillary electrophoresis as the primary method - HPLC is now the gold standard for both screening and quantification 3
• Alkaline pH electrophoresis (pH 8.6) serves as initial screening, but requires confirmatory testing 3
• Electrophoresis alone is insufficient - some variants are electroforetically silent and will be missed 4, 3

Confirmation Algorithm

• Follow positive screening with acid electrophoresis (pH 7.1 or citrate agar) since different hemoglobins may co-migrate at alkaline pH 3, 5
• Perform targeted DNA sequencing of α- and β-globin genes for definitive identification 4, 3
• Measure methemoglobin levels spectrophotometrically (normal <1%) when HbM variants are suspected 3

Specific Clinical Scenarios

Transfusion Complications in Sickle Cell Disease

• Order hemoglobin electrophoresis when patients with known sickle cell disease develop severe hemolysis post-transfusion 6
• Complete absence of hemoglobin A on electrophoresis during hemolytic event indicates severe delayed hemolytic transfusion reaction (DHTR) rather than hyperhemolysis syndrome 6
• This distinction is critical because hyperhemolysis has worse prognosis and higher recurrence rates with subsequent transfusions 6

Prenatal Diagnosis

• Hemoglobin electrophoresis of fetal hemolysate provides a simple, rapid alternative to PCR for prenatal exclusion of sickle cell anemia 7
• Particularly useful when rapid results are needed due to advanced gestational age 7
• Genetic testing enables prevention of thalassemia births and pregnancy complications 8

Obstetric Patients

• Women with known sickle cell disease should be managed in specialized obstetric units given 1-3% maternal mortality and 1-8% perinatal mortality rates 1
• High incidence of complications: 57% painful crises, 38% cesarean delivery, 23% ICU admission 1

Critical Pitfalls to Avoid

• Never skip family studies when hemoglobinopathy is identified - this is essential for genetic counseling and family planning 4
• Do not interpret HbA1c in patients with hemoglobinopathies unless using assays without interference (check www.ngsp.org/interf.asp) 3
• Avoid ordering in neonates or recently transfused patients if using solubility testing, as false negatives occur 2
• Do not order for routine anemia workup without first evaluating iron studies and red cell indices 1
• Always clearly communicate suspected diagnosis on laboratory request forms to ensure appropriate processing and interpretation 2