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 ★ 85

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

"Shows proper gel setup and running of acrylamide gels, helping avoid uneven polymerization issues."

Bilibili (China-Accessible Mirrors) ★ 70

Western Blot Complete Process First-Person View

"First-person Western blot protocol demonstrates correct gel casting and handling steps."

Cell Signaling Technology ★ 70

Western Blotting Protocol

"Detailed Western blot protocol covers gel preparation and electrophoresis, useful for troubleshooting band distortion."