Failure Case Library

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

All Techniques (7) 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)
HMW DNA Extraction (Monarch) severe

Low DNA Yield from HMW Extraction

Recovered DNA concentration is significantly lower than expected based on input amount. Spectrophotometric measurements may show unexpectedly low concentrations for UHMW DNA samples.

💡 6 causes ✓ 6 fixes
HMW DNA Extraction (Monarch) severe

Cell or Leukocyte Loss During Pelleting

Lower than expected DNA yield despite using correct input amounts. Cell pellet may not be visible when working with low cell numbers or certain frozen blood samples.

💡 4 causes ✓ 4 fixes
HMW DNA Extraction (Monarch) critical

DNA Fragmentation and Size Reduction

Extracted DNA shows reduced fragment size on gel electrophoresis or pulse-field gel analysis. UHMW DNA appears degraded with smaller molecular weight distribution than expected.

💡 6 causes ✓ 6 fixes
HMW DNA Extraction (Monarch) moderate

Eluted DNA Difficult to Dissolve or Resuspend

DNA pellet or eluate shows poor solubility with visible aggregates or turbidity. DNA remains undissolved after standard elution protocol, requiring extended incubation.

💡 5 causes ✓ 6 fixes
HMW DNA Extraction (Monarch) severe

Protein Contamination in Purified DNA

Purified DNA shows poor A260/A230 ratio or visible turbidity. Solution may appear cloudy due to protein contamination, affecting downstream applications.

💡 6 causes ✓ 6 fixes
HMW DNA Extraction (Monarch) severe

Low Yield from Tissue Samples

DNA recovery from tissue significantly lower than expected, particularly from muscle, brain, or metabolically active organs. Upper phase transfer yields less DNA than anticipated.

💡 5 causes ✓ 5 fixes
HMW DNA Extraction (Monarch) moderate

Inaccurate Cell Count Leading to Protocol Mismatch

Unexpectedly high or low DNA yield that does not correlate with estimated cell count. Certain cultured cells appear clumped making accurate counting difficult.

💡 4 causes ✓ 5 fixes