Failure Case Library

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

All Techniques (9) 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)
ELISA Development moderate

Signal Drift Across Plate

Gradual change in signal intensity across the plate suggesting time-dependent variation during assay setup

💡 2 causes ✓ 2 fixes
ELISA Development minor

Edge Effects on Plate

Wells at the edge of the plate show different readings compared to central wells

💡 1 causes ✓ 2 fixes
ELISA Development moderate

Very Low Readings Across Plate

Consistently low optical density readings across entire plate including standards

💡 5 causes ✓ 6 fixes
ELISA Development severe

Poor Assay-to-Assay Reproducibility

Inconsistent results between different assay runs with same samples

💡 7 causes ✓ 9 fixes
ELISA Development moderate

Poor Duplicate Reproducibility

Duplicate wells showing inconsistent results with high variation between replicates

💡 6 causes ✓ 8 fixes
ELISA Development moderate

Poor Standard Curve Discrimination

Standard curve achieved but shows low or flat curve with poor discrimination between concentration points

💡 6 causes ✓ 8 fixes
ELISA Development severe

Excessive Signal - Entire Plate Turned Blue

Whole plate turned uniformly blue indicating non-specific signal saturation

💡 5 causes ✓ 5 fixes
ELISA Development critical

Complete Signal Absence

No detectable signal across the entire plate when signal is expected

💡 7 causes ✓ 9 fixes
ELISA Development moderate

High Background Signal

Elevated background noise across the ELISA plate making it difficult to distinguish specific signals from non-specific binding

💡 3 causes ✓ 3 fixes