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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 (2)
Cell Signaling Technology ★ 85
Western Blot Troubleshooting Guide
"Dedicated troubleshooting guide directly addresses diagnostic methods for weak/no signal problems in Western blots, the exact symptom of the failure case."
Bilibili (China-Accessible Mirrors) ★ 72
Western blot full protocol: Protein extraction to chemiluminescence
"Complete hands-on workflow including chemiluminescence detection step where sodium azide interference with peroxidase reactions would manifest as signal loss."
Source: sigmaaldrich.com ↗
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