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)
End-point PCR Primers moderate

Primers Stop Working Over Time

Primers that initially worked successfully for PCR gradually fail or produce poor results after storage

💡 4 causes ✓ 5 fixes
End-point PCR Primers severe

Subcloning Failure with Restriction Site Primers

Very few or no colonies obtained when subcloning PCR products generated with primers containing restriction enzyme sites, despite successful amplification

💡 4 causes ✓ 5 fixes
End-point PCR Primers severe

GC-Rich Template Amplification Failure

PCR fails to amplify templates with high GC content or produces very low yields

💡 5 causes ✓ 8 fixes
End-point PCR Primers minor

High Molecular Weight Material Stuck in Wells

Ethidium bromide-stainable material remains in the gel wells and does not migrate during electrophoresis

💡 3 causes ✓ 4 fixes
End-point PCR Primers severe

Unexpected PCR Product

PCR produces a band of incorrect size or sequence not matching the expected target

💡 5 causes ✓ 6 fixes
End-point PCR Primers critical

No PCR Product (Complete Failure)

No bands visible on gel electrophoresis after PCR amplification

💡 10 causes ✓ 10 fixes
End-point PCR Primers severe

Low PCR Product Yield

Desired PCR fragment appears faint or barely visible on gel electrophoresis

💡 14 causes ✓ 14 fixes
End-point PCR Primers moderate

PCR Product Smearing on Gel

Gel electrophoresis shows smeared or streaked bands instead of distinct sharp bands

💡 6 causes ✓ 7 fixes
End-point PCR Primers moderate

Nonspecific Bands in PCR

Multiple unwanted bands appear on gel electrophoresis alongside or instead of the desired PCR product

💡 4 causes ✓ 5 fixes