Low IP Efficiency from Incorrect Bead Selection (Immunoprecipitation (CST Guide))

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Weak IP signal despite good input control. Low recovery of target protein compared to expected levels based on antibody quality. Common causes: 1) Protein G beads used with rabbit antibody instead of higher-affinity Protein A beads 2) Protein A beads used with mouse antibody instead of higher-affinity Protein G beads 3) Suboptimal IgG-bead binding due to species-bead mismatch 4) Insufficient bead capacity or binding time for antibody-antigen complex capture Solutions: 1) Use Protein A beads when performing IP with rabbit antibodies (higher affinity for rabbit IgG) 2) Use Protein G beads when performing IP with mouse antibodies (higher affinity for mouse IgG) 3) Consider combination Protein A/G beads to increase IgG binding efficiency 4) Optimize bead volume (typically 20-40 µL bead slurry) and incubation time (1-4 hours at 4°C)