Low/No IP Signal from Insufficient Target Expression

Symptom

IP experiment yields no detectable signal on western blot. Input lysate control shows weak or absent target protein bands, indicating expression below detection threshold.

1. 1 Target protein or interacting proteins expressed at levels below western blot detection limit
2. 2 Cell line or tissue model does not sufficiently express the target protein of interest
3. 3 Inappropriate cell line or tissue selection based on expression profiling data

1. 1 Use expression profiling tools (BioGPS, The Human Protein Atlas) and literature to verify target expression in chosen model
2. 2 Include known positive control from Control Treatments by Target table to confirm experimental setup
3. 3 Include input lysate control to verify target protein expression levels and antibody reactivity
4. 4 Select alternative cell lines or tissues with documented higher expression of target protein

Related Video (2)

Cell Signaling Technology ★ 95

Western Blotting Protocol

"Directly demonstrates western blot protocol, the detection method used to identify the low/no signal problem in this IP failure case"

Cell Signaling Technology ★ 78

How CUT&RUN Profiles Chromatin | Cell Signaling Technology

"CUT&RUN is explicitly presented as a low cell number alternative to chromatin IP, providing context for when standard IP fails due to insufficient target expression"