Uneven Bands Due to Improper Gel Polymerization

Symptom

Bands appear distorted, wonky, or uneven across lanes. In extreme cases, the same protein appears at different molecular weights in different lanes due to irregular gel matrix formation.

1. 1 Incorrect TEMED concentration in gel recipe causing incomplete or uneven polymerization
2. 2 Gel not fully covered with buffer during setting process
3. 3 Uneven polymerization across gel thickness or width
4. 4 Temperature fluctuations during gel polymerization phase

1. 1 Verify gel recipe and confirm correct TEMED volume is added
2. 2 Ensure gel is completely covered with buffer during polymerization
3. 3 Allow adequate polymerization time (minimum 30-45 minutes for resolving gel)
4. 4 Prepare fresh acrylamide solutions and check ammonium persulfate (APS) activity

Related Video (3)

Bio-Rad Laboratories ★ 78

Setting Up and Running Mini-PROTEAN® TGX™ Precast Gels

"Demonstrates proper setup and running of acrylamide gels, the exact system where TEMED-driven polymerization matters."

Cell Signaling Technology ★ 72

Western Blotting Protocol

"Walks through the full Western blot protocol with emphasis on reproducible results, helping avoid gel-related band artifacts."

Bilibili (China-Accessible Mirrors) ★ 70

Western blot hands-on: Protein extraction through detection

"Hands-on Western blot protocol that explicitly covers troubleshooting common pitfalls, including electrophoresis problems."