Serum protein electrophoresis

Principle

• At alkaline pH (about 8.6, barbital buffer) serum proteins carry net negative charge and migrate toward the anode in an electric field.
• Separation depends on charge, size and shape; support media are agarose gel or cellulose acetate, or capillary zone electrophoresis in automated systems.
• After fixing and staining (Ponceau S, amido black or Coomassie blue), bands are quantified by densitometry.

Specimen

• Serum is required. Plasma contains fibrinogen, which forms a band between β and γ that can mimic a monoclonal band.
• Avoid haemolysis: haemoglobin–haptoglobin complexes appear in the α2/β region.

Results

• Five main zones from anode: albumin (largest band), α1 (mainly α1-antitrypsin), α2 (α2-macroglobulin, haptoglobin), β (transferrin, C3; may split into β1 and β2), γ (immunoglobulins).
• Typical serum albumin 3.5–5.0 g/dL; local reference ranges apply.

Interpretation

• Monoclonal gammopathy: narrow, dense M-spike, usually in γ or β region (multiple myeloma, MGUS, Waldenström). Confirm and type by immunofixation.
• Polyclonal increase: broad diffuse γ increase (chronic infection, autoimmune disease, chronic liver disease).
• Cirrhosis: low albumin with β–γ bridging (IgA increase).
• Acute inflammation: raised α1 and α2 bands.
• Nephrotic syndrome: low albumin and γ, markedly raised α2 (α2-macroglobulin retained).
• α1-antitrypsin deficiency: faint or absent α1 band.
• Hypogammaglobulinaemia: pale γ zone; can also be due to light-chain myeloma.

Quality control and pitfalls

• Run a normal control serum on each gel; check band separation and staining.
• Small M-proteins may hide within β band; free light chains may not show on SPE.

Clinical use

• Screening for plasma cell disorders and evaluating protein abnormalities in liver, renal and inflammatory disease.