Serum protein is separated in 1% agarose gel, 25minutes at 100 volts, stained with Ponseau S protein stain. The
separation of the protein is not optimal. What would you do to improve the separation

(a) Increase the agarose concentration
(b) Run the electrophoresis at 75V instead of 100
(c) Run the electrophoresis for 15 minutes instead of 25 minutes
(d) Use a lipid stain instead of protein stain

Correct answer: (A) Increase the agarose concentration

Explanation:

The resolution of protein separation in agarose gel electrophoresis depends on several factors, including gel concentration, voltage, and running time.

• Increasing the agarose concentration improves the pore size of the gel, allowing better separation of serum proteins based on charge and size.
• Serum proteins vary in molecular weight, and using a higher agarose concentration (e.g., 1.5% or 2% instead of 1%) provides better resolution, especially for smaller proteins.

Why Not the Other Options?

• (B) Run the electrophoresis at 75V instead of 100V ❌

  • Lower voltage slows down migration and may reduce resolution instead of improving it.
  • Optimizing agarose concentration is more effective than lowering voltage.

• (C) Run the electrophoresis for 15 minutes instead of 25 minutes ❌

  • Shortening the run time may lead to insufficient separation, causing poor resolution.
  • Instead, running for a longer time at optimal voltage could enhance separation.

• (D) Use a lipid stain instead of a protein stain ❌

  • Ponseau S is a protein stain, which is appropriate for serum protein analysis.
  • A lipid stain (e.g., Sudan Black) is not relevant for protein separation.

Final Answer:

✅ (A) Increase the agarose concentration