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)
Cell Culture (Cell Death) moderate

Replicative Senescence After Excessive Passaging

Primary or finite cell lines show progressive slowing of growth, enlarged flattened morphology, failure to reach confluence, and increased spontaneous death after many passages.

💡 4 causes ✓ 5 fixes
Cell Culture (Cell Death) severe

Cell Death from CO₂/pH Imbalance in Bicarbonate Buffer

Cells show progressive death over 24-48 hours with media color shift (phenol red turning yellow or purple). Cells exhibit membrane damage and eventual lysis despite sterile conditions.

💡 5 causes ✓ 5 fixes
Cell Culture (Cell Death) moderate

Photooxidative Cell Death from Fluorescent Light Exposure

Cells cultured near fluorescent lights or windows show increased death rates, membrane blebbing, and oxidative stress markers. HEPES-buffered media particularly affected.

💡 4 causes ✓ 5 fixes
Cell Culture (Cell Death) moderate

Contact Inhibition and Nutrient Depletion Cell Death

Cultures exceeding 80-100% confluence show central necrosis, detachment of cell sheets, and debris accumulation. Media becomes acidic (yellow) and depleted.

💡 4 causes ✓ 5 fixes
Cell Culture (Cell Death) severe

Cell Death from Poor Quality or Toxic Reagents

Sudden cell death occurs immediately after introducing new lot of media, serum, trypsin, or buffer. Previously healthy cultures show rapid necrosis or apoptosis within 12-24 hours.

💡 5 causes ✓ 6 fixes
Cell Culture (Cell Death) critical

Cell Death from Bacterial or Fungal Contamination

Media becomes rapidly turbid with visible particles or cloudiness. Cells show signs of toxicity, detachment, and death. Microscopy reveals motile bacteria or fungal filaments. Note: mycoplasma rarely causes acute cell death.

💡 4 causes ✓ 6 fixes
Cell Culture (Cell Death) critical

Cell Death Due to Incubator Temperature Instability

Previously healthy cultures show sudden widespread cell death with membrane blebbing, crenation, and cellular debris. Cell death occurs within hours of incubation.

💡 4 causes ✓ 5 fixes
Cell Culture (Cell Death) critical

No/Few Viable Cells After Thawing Cryopreserved Stock

Upon thawing cryopreserved cell stocks, researchers observe minimal or no viable cells via trypan blue exclusion or hemocytometer counting. Cells appear non-adherent, crenated, or lysed.

💡 5 causes ✓ 5 fixes