ArticleBMC research notes2025
Optimization of a thermochemical antibody stripping method for enhanced tyramide signal amplification-based opal multiplex immunohistochemistry in fragile tissues.
Article in BMC research notes, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 1 paper.
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Who cites it
1 citing paper in PubMed.
- Volumetric Cyclic Immunofluorescence for 3D Spatial Profiling of Immune Structures in Human FFPE Tissue.bioRxiv : the preprint server for biology · 2026Article
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Authors and funding
6 authors.
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Abstract
objectiveTyramide signal amplification (TSA)-induced Opal multiplex immunohistochemistry (mIHC) represents an advanced methodology for the in-situ detection of multiple target proteins. A critical aspect of this technique is the complete removal of primary and secondary antibodies to prevent signal cross-reaction. However, current antibody stripping methods exhibit several limitations, particularly when applied to tissues prone to delamination. Microwave treatment may compromise tissue integrity, additionally, the effectiveness of chemical reagents can be sensitive to variations in temperature, pH, and concentration. This study aims to optimize antibody stripping strategies specifically for use in Opal mIHC protocols, addressing limitations of current methods which particularly for tissues prone to delamination.
resultsEvaluation of microwave oven-assisted antibody removal (MO-AR), chemical reagent-based antibody removal (CR-AR), and hybridization oven-based stripping at 50 °C (HO-AR-50) and 98 °C (HO-AR-98) revealed that MO-AR and HO-AR-98 most effectively removed primary and secondary antibodies. In five-color mIHC on mouse kidney sections, both methods yielded strong target-specific signals. However, in brain tissue sections prone to delamination, HO-AR-98 better preserved tissue integrity compared to MO-AR and supported effective multi-target staining. These findings establish a novel thermochemical stripping method compatible with TSA-based Opal mIHC, enhancing its utility in research and clinical diagnostics.
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