Antibody not validated for IHC application or tissue type

Symptom

Complete absence of staining with antibody that works in other applications. The antibody may not recognize native protein conformation present in tissue sections.

1. 1 Antibody designed for denatured proteins (western blot) not recognizing native epitopes in IHC
2. 2 Antibody not validated for specific tissue preparation method (FFPE vs. fresh frozen)
3. 3 Epitope only accessible in denatured state, not in tissue architecture
4. 4 Antibody specificity incompatible with IHC application requirements

1. 1 Check antibody datasheet to confirm IHC validation
2. 2 Verify antibody is validated for specific tissue preparation (FFPE, fresh frozen, etc.)
3. 3 Select antibodies with confirmed reactivity in native protein state
4. 4 Consider alternative antibodies specifically validated for IHC with your tissue type
5. 5 Test antibody in pilot experiment with known positive control tissue

Related Video (3)

Cell Signaling Technology ★ 85

Tissue Controls & More for Immunohistochemistry | CST Tech Tips

"Directly addresses how to verify antibody validation and use controls to troubleshoot absent staining in IHC."

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Immunohistochemistry on Paraffin-Embedded Sections (CST Demo)

"Shows proper IHC protocol on paraffin sections, helping confirm procedural steps when troubleshooting staining failure."

Cell Signaling Technology ★ 70

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"Focuses on antigen retrieval, a key step affecting epitope availability critical for antibody recognition in tissue."