Order of Migration in Serum Protein Electrophoresis
In serum protein electrophoresis, the correct order of migration from cathode to anode is (-) gamma, beta, alpha-2, alpha-1, albumin (+).
Principles of Serum Protein Electrophoresis
Serum protein electrophoresis (SPEP) is a laboratory technique that separates serum proteins based on their electrical charge-to-mass ratio in an alkaline buffer (typically pH 8.6) 1. During this process:
- Proteins migrate toward the electrode of opposite charge
- The cathode is negatively charged (-)
- The anode is positively charged (+)
- Proteins with greater negative charge migrate faster toward the anode
Migration Pattern Explanation
The migration pattern follows this specific order due to the inherent electrical properties of each protein fraction:
- Gamma globulins: Migrate least toward the anode (remain closest to cathode) due to their relatively neutral charge
- Beta globulins: Migrate slightly further than gamma globulins
- Alpha-2 globulins: Migrate further than beta globulins
- Alpha-1 globulins: Migrate further than alpha-2 globulins
- Albumin: Migrates furthest toward the anode due to its strong negative charge at alkaline pH
Technical Considerations
Several factors influence the quality and reliability of SPEP results:
- Buffer pH: The optimal pH for serum protein electrophoresis is 8.6, which allows for proper separation of the protein fractions 1
- Temperature control: Maintaining consistent temperature during electrophoresis is essential as it affects protein migration patterns 2
- Electrophoresis duration: The extent of protein migration is directly proportional to the duration of electrophoresis 2
- Voltage gradient: The strength of the electrophoretic field directly affects migration distance 2
Clinical Applications
SPEP is a critical diagnostic tool used for:
- Detection of monoclonal gammopathies
- Evaluation of multiple myeloma and related disorders 2
- Assessment of protein abnormalities in various disease states
- Monitoring patients with plasma cell disorders
Common Pitfalls to Avoid
- Misidentification of fractions: The beta-globulin fraction can sometimes be misidentified, particularly when C3 complement appears in different positions 3
- Interference factors: Several substances can cause artifacts in SPEP results, including lipid emulsions, hemoglobin, contrast agents, and certain plasma substitutes 4
- Quantification limitations: There is greater imprecision and reduced accuracy when measuring low concentration M-proteins 5
- Migration shifts: Some proteins may show gamma-to-beta migration shifts, affecting proper identification 4
Understanding the correct order of migration is fundamental for accurate interpretation of SPEP patterns and proper diagnosis of protein-related disorders.