Failure Case Library

Real lab failures, root causes, and fixes — curated and bilingually annotated by our team.

All Techniques (8) PCR (Sigma Guide) (12) End-point PCR Primers (9) Western Blot (9) Wound Healing Assay (5) Cell Culture (Contamination) (7) Lipid Transfection (8) CCK-8 Cell Viability Assay (5) Colony Formation Assay (10) Transfection (8) Nucleic Acid Quantification (9) Transfection (Co-transfection) (1) Protein Extraction (4) Plasmid Construction (Double Digest) (1) Sanger Sequencing (2) Plasmid Construction (1) qPCR (RT-qPCR) (5) Transwell Migration / Invasion Assay (5) Immunohistochemistry (IHC) (6) PCR (Polymerase Chain Reaction) (26) Restriction Enzyme Digest (13) DNA Cleanup & Plasmid Purification (7) RNA Cleanup (4) NGS Library Preparation (NEBNext Ultra II) (7) HMW DNA Extraction (Monarch) (7) LAMP (Loop-mediated Isothermal Amplification) (7) RNA Depletion for RNA-seq (7) Bacterial rRNA Depletion (4) Cell-free DNA Extraction (8) ELISA (Competitive) (18) ELISA (Signal Problems) (11) ELISA (High Background) (8) ELISA (Inconsistent Results / High CV) (6) ELISA (Standard Curve Fit Problems) (6) Western Blot (Weak / No Signal) (6) Western Blot (Detection Problems) (7) Western Blot (Bands at Wrong MW) (5) Western Blot (Misshapen / Uneven Bands) (5) Western Blot (Unexpected Multiple Bands) (7) Western Blot (Unusual Gel Band Appearance) (3) ChIP (High Background) (1) ChIP (Low Resolution with High Background) (6) ChIP (Low Signal) (8) ChIP (PCR Amplification Problems) (4) Immunohistochemistry (High Background) (9) Immunohistochemistry (No Staining) (9) Immunoprecipitation (High Antibody Elution) (1) Immunoprecipitation (High Background) (8) Immunoprecipitation (No Protein Detected) (5) Immunoprecipitation (Protein Obstruction) (1) ELISPOT (8) Tissue Imaging (Autofluorescence) (9) Flow Cytometry (Troubleshooting) (8) ELISA Development (9) Flow Cytometry (Autofluorescence) (7) Flow Cytometry (Compensation) (7) Flow Cytometry (Fc Blocking) (7) Flow Cytometry (Fixation & Permeabilization) (9) Flow Cytometry (Isotype Controls) (7) Flow Cytometry (Fixation Buffers) (7) Western Blot (CST Guide) (8) ChIP (CST Guide) (8) Immunoprecipitation (CST Guide) (14) Immunohistochemistry (CST Guide) (14) Flow Cytometry (CST Guide) (8) ELISA (R&D Guide) (10) ELISA (Sigma Guide) (6) Western Blot Immunodetection (19) IP-Western Blot (6) Flow Cytometry (Sample Considerations) (14) Flow Cytometry (Paraformaldehyde Fixation) (14) Western Blot (Sigma Protocol) (8) PCR / RT-PCR Amplification Problems (5) Cell Culture (Cell Death) (8) Cell Culture (Precipitates) (6) PCR (Invitrogen Guide) (8) PCR / qPCR Plastics (9) Flow Cytometry (Controls) (8) Plasmid Mini-prep (1) Transfection (siRNA Knockdown) (1) Western Blot (Blue Background) (1)
ChIP (Low Signal) severe

Low X-ChIP Signal from Excessive Formaldehyde Cross-Linking

Low signal specifically in X-ChIP (cross-linked ChIP) experiments. Antibody appears unable to bind target epitopes despite proper antibody concentration and chromatin quality.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) moderate

Loss of Specific Signal Due to Overly Stringent Wash Conditions

Low signal at expected target regions while background is also very low. Positive control regions show reduced signal compared to expected levels.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) moderate

Low ChIP Signal from Ineffective Cell Lysis

Overall low signal with visible cell clumps or debris in lysate. Chromatin yield is lower than expected based on starting cell number.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) minor

No Signal at Region of Interest Due to Absent Target

No signal detected at the specific region of interest while ChIP procedure appears technically successful. Other genomic regions or positive controls may show expected signals.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) severe

Low Signal from Using N-ChIP for Weak DNA-Binding Proteins

Consistently low or absent signal when studying transcription factors or chromatin-associated proteins using native ChIP (N-ChIP). Histone ChIP experiments work well in the same laboratory.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) severe

Low ChIP Signal Due to Chromatin Over-Fragmentation

Low or absent signal in ChIP experiment despite proper antibody and starting material. Chromatin fragments are smaller than 500 bp after sonication or enzymatic digestion.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) moderate

Low ChIP Signal from Insufficient Starting Material or Antibody

Weak or barely detectable signal across all samples including positive controls. Signal intensity is uniformly low rather than selectively absent at specific regions.

💡 4 causes ✓ 4 fixes
ChIP (Low Signal) severe

Low Recovery Due to Incompatible Antibody Affinity Beads

Low signal across all samples with high background. Antibody appears present in supernatant after IP, suggesting poor capture by beads.

💡 4 causes ✓ 5 fixes