Electrophoresis of Serum Proteins Buffer pH
The most commonly used buffer pH for electrophoresis of serum proteins is 8.6 (option C). 1, 2
Scientific Basis for pH 8.6 in Serum Protein Electrophoresis
Serum protein electrophoresis is traditionally performed at an alkaline pH of 8.6 for several important reasons:
- At pH 8.6, most serum proteins are negatively charged, allowing for optimal separation based on their charge-to-mass ratio
- This specific pH provides the best resolution between the major serum protein fractions (albumin, alpha-1, alpha-2, beta, and gamma globulins)
- The standard buffer used historically is 5'-diethylbarbiturate (barbital) at pH 8.6 1
Buffer Composition Considerations
When performing serum protein electrophoresis, several factors influence the choice of buffer:
- Traditional systems: Sodium barbital electrophoresis buffer at pH 8.6 is widely used in agarose gel systems 2
- Supporting media: Tris-EDTA-borate-buffered agarose gels (pH 8.8) are often paired with barbital buffer (pH 8.6) 2
- Capillary electrophoresis: While traditional barbital buffers have high UV absorbance limiting their use in capillary systems with UV detection, alternative buffers maintaining the same pH range are used 1
Technical Considerations
The pH of the buffer significantly impacts separation quality:
- At pH 8.6, the optimal balance between protein charge and stability is achieved
- Lower pH values (options A and B) would result in poor separation of serum proteins as many would be closer to their isoelectric points
- Higher pH values (option D) could cause protein denaturation or excessive negative charge, reducing separation resolution
Practical Applications
In clinical laboratories, serum protein electrophoresis at pH 8.6 is routinely used for:
- Detection of monoclonal gammopathies
- Evaluation of protein abnormalities in various disease states
- Monitoring patients with multiple myeloma and other plasma cell disorders
Quality Control Considerations
When performing electrophoresis of serum proteins:
- Buffer pH should be verified prior to starting electrophoresis as it significantly affects migration patterns 3
- Fresh buffers should be prepared to ensure consistent pH and reliable results
- Temperature control during electrophoresis is essential as it can affect protein migration 3
The consistent use of pH 8.6 buffer systems across decades of clinical practice has established it as the standard for serum protein electrophoresis, providing reliable and reproducible separation of serum protein fractions.