No Protein Detected: Insufficient Antibody Capture (Immunoprecipitation (No Protein Detected))

Question

Accepted Answer

No target protein is detected in the elution fraction after immunoprecipitation procedure is completed. Western blot or other detection methods show no signal for the target protein. Common causes: 1) Antibody concentration is below the minimum required for effective capture of target protein 2) Insufficient antibody amount used relative to the abundance of target protein in sample 3) Antibody binding capacity exceeded by excess target protein without proportional antibody increase Solutions: 1) Check vendor datasheet for recommended antibody amount per IP reaction and verify current usage meets minimum 2) Titrate antibody concentration upward in 2-fold increments (e.g., 2 µg, 4 µg, 8 µg) to optimize capture efficiency 3) Calculate antibody:protein molar ratio and adjust antibody amount proportionally to sample protein concentration