What is Tyramide Signal Amplification (TSA)?

Tyramide signal amplification (TSA) is a novel technique designed to enhance the detection of low-abundance targets (e.g. proteins and nucleic acid sequences). It can be readily integrated into any application that utilizes horseradish peroxidase (HRP) in its protocol. These applications include ELISA, immunocytochemistry (ICC), immunohistochemistry (IHC) and in situ hybridization (FISH). Advantages of tyramide signal amplification, include:

• Detection of low-abundance targets
• Higher precision and sensitivity up to 100-fold that of conventional ICC, IHC and FISH methods
• Easy integration with ICC, IHC and FISH applications
• Conservation of primary and secondary antibodies

TSA Principle & Workflow

TSA - sometimes called Catalyzed Reporter Deposition or CARD, is an enyzme-mediated detection method that utilizes the catalytic activity of HRP for the covalent deposition of labeled tyramide on and near target proteins or nucleic acid sequences in situ. In the presence of hydrogen peroxide (H₂O₂), HRP converts labeled tyramide substrates into highly-reactive, short-lived tyramide radicals that rapidly bind to tyrosine residues on and proximal to the enzyme site. These labels can be detected by standard chromogenic or fluorescent techniques.

TSA labeling procedure consists of the following steps:

1. Sample preparation – fixation and permeabilization of cells or tissue sample
2. Blocking of nonspecific epitopes
3. Binding with primary antibody, biotinylated primary antibody or biotinylated nucleic acid probe
4. Binding with HRP-secondary antibody or HRP-streptavidin
5. Incubation with tyramide substrate
6. Mount sample and image signal