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Western Blot (Sigma Protocol) moderate

Enzyme Inhibitor Interference

Symptom
No signal or very weak signal despite proper antibody binding
Common Causes
  1. 1 Sodium azide present in buffers inhibiting peroxidase reactions
  2. 2 Over-washing removing enzyme conjugates
Solutions
  1. 1 Do not use sodium azide in buffers when using peroxidase-conjugated antibodies
  2. 2 Shorten wash times and omit detergents from washing buffers
Related Video (3)
Cell Signaling Technology ★ 90
Western Blot Troubleshooting Guide
"Western blot troubleshooting guide directly addresses diagnosing weak/no signal issues like enzyme inhibitor interference."
Bilibili (China-Accessible Mirrors) ★ 72
Western blot hands-on: Protein extraction through detection
"Hands-on protocol with emphasis on troubleshooting common pitfalls, including buffer/reagent issues affecting detection."
Bilibili (China-Accessible Mirrors) ★ 70
Reliable and Reproducible Western Blot Results
"CST webinar on reliable results covers proper buffer preparation and reagent pitfalls that can cause signal failure."
Source: sigmaaldrich.com ↗
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