Failure Case Library

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

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

Bands Appear Lower Than Expected Molecular Weight

All protein bands or the target band migrate further down the membrane than expected based on the predicted molecular weight. Coomassie blue staining of the membrane confirms abnormally low band positions before immunostaining.

💡 4 causes ✓ 4 fixes
Western Blot (Bands at Wrong MW) moderate

Low Acrylamide Percentage for Target Protein Size

Lower molecular weight proteins migrate excessively fast through the gel, resulting in poor resolution and bands appearing at the bottom of the membrane or running off entirely.

💡 3 causes ✓ 4 fixes
Western Blot (Bands at Wrong MW) moderate

Electrophoresis Time Not Optimized for Target Protein

Protein bands do not align with expected molecular weight markers, with systematic upward or downward shifts affecting all bands uniformly. Pre-staining reveals migration issues before antibody detection.

💡 4 causes ✓ 5 fixes
Western Blot (Bands at Wrong MW) moderate

Bands Appear Higher Than Expected Molecular Weight

All protein bands or the target band remain near the top of the membrane, migrating less than expected based on the predicted molecular weight. Coomassie blue staining confirms abnormally high band positions before immunostaining.

💡 4 causes ✓ 4 fixes
Western Blot (Bands at Wrong MW) moderate

High Acrylamide Percentage for Target Protein Size

Higher molecular weight proteins remain near the loading wells with minimal migration, resulting in poor resolution and bands clustered at the top of the membrane.

💡 4 causes ✓ 4 fixes