Tetraethylammonium salt signal amplification (TSA, Tyramide signal amplification) is a class of enzyme-based detection methods that use horseradish peroxidase (HRP) to label target proteins. Similar to the conventional DAB colorimetric method in immunohistochemistry, TSA also employs HRP-labeled secondary antibodies and has a corresponding "color development" step (HRP catalyzes the addition of tyramine fluorescein substrate to the reaction system, producing an active fluorescent substrate. This active substrate can covalently bind with tyrosine residues or other residues on the antigen, forming stable covalent conjugates of tyramine fluorescein on the sample. Subsequently, non-covalently bound primary antibody-secondary antibody-HRP complexes are washed away using thermal repair or antibody elution buffer. The process then repeats with another primary antibody-HRP secondary antibody for a second round of incubation, followed by a different tyramine fluorescein substrate. This cycle continues to achieve multiplex labeling. It is primarily used in immunohistochemical (IHC), immunofluorescent (IF), or in situ hybridization (ISH) experiments.

The core principle is:

HRP catalytic reaction: after the primary antibody binds to the target molecule, the secondary antibody coupled with horseradish peroxidase (HRP) binds to the primary antibody and activates the subsequent reaction.

Tyramine substrate deposition: HRP catalyzes the addition of tyramine substrates (such as fluorescent or chromogenic tyramine molecules) to generate highly reactive free radicals, which covalently bind to protein tyrosine residues near the antigen to form stable signal deposition.

Signal cascade amplification: A single HRP enzyme can catalyze the deposition of a large number of tyramine molecules in a short period of time, significantly amplifying the target signal and enabling the detection of low abundance targets.

Product superiority:

• Ultra-high sensitivity: through enzyme catalytic cascade reaction, the signal intensity can be increased by 10-100 times, suitable for trace protein or nucleic acid detection.
• Multi-channel compatibility: Supports multi-color labeling (such as tyramine dye with different fluorescence wavelengths) to achieve multiple target analysis in the same sample.
• Low background interference: the covalently bound tyramine signal is highly localized, reducing non-specific adsorption and improving SNR.
• Wide applicability: compatible with paraffin sections, frozen sections, cell crawling sheets and whole tissue transparent samples (such as organoids, 3D culture models).
• Flexible detection mode: provides two detection options of fluorescence (such as FITC, Cy3, Cy5) and color (such as DAB), suitable for ordinary microscope or high resolution confocal imaging system.
• Instantaneous design: pre-optimized reagent formula, simplified operation process, shortened experimental time.

Application scenarios:

• Low abundance target detection: such as rare cell markers, synaptic proteins, cytokines in the tumor microenvironment.
• Multiple marker analysis: multiple protein or nucleic acid targets are detected simultaneously in a single sample (requires multiple rounds of antibody stripping and regeneration).
• Enhanced in situ hybridization: improve the sensitivity of in situ detection of RNA (such as miRNA, lncRNA) or DNA.
• 3d imaging support: deep imaging is suitable for thick tissue sections or transparent samples to enhance signal penetration.
• Diagnosis and drug development: validation of biomarkers for pathological diagnosis, drug target location and efficacy evaluation.
• Optimization of in vivo imaging: improve the real-time detection efficiency of in vivo or dynamic samples by signal amplification.

Our TSA kit has a complete set of components: Fluorescent Dye, TSA buffer, Secondary Antibody, DAPI Stainin solution, Antibody Diluention, 3% Hydrogen Peroxide and Anti fluorescence quenching sealing agent.

List of kit products:

For more information please email us info@kj-bio.cn.