Tyramine signal amplification

Tyramine signal amplification (TSA) reagent, for example, TSA-FITC (PerkinElmer). 

Different authors and commercial suppliers have assigned different names to signal amplification using tyramine. For example, tyramine signal amplification (TSA) system and the catalyzed signal amplification (CSA) system are comma-daily available from DuPont NEN Life Science Products, Boston, MA, and DAKO Corporation, Carpinteria, CA, respectively. In addition, the tarns CARD (catalyzed reporter deposition) (Bobrow et al., 1989), TA (tyramide amplification) (Shindler and Roth, 1996), and ImmunoMax (Merz et al., 1995) have been used for the tyramine amplification technique. The use of different names for almost identical procedures has resulted in confusion. To standardize the terminology, the neutral abbreviation, tyramide amplification technique (TAT) should be accepted (Von Wasielewski et al., 1997). 

Fig. 6.2 Tyramide signal amplification. T is the labeled tyramine and HRP, horse radish peroxidase. The Label can be a lluorochrome or biotin. The lluorochrome can be visualized directly in a fluorescence microscope. Biotin can be visualized via labeled streptavidin...

Tyramide signal amplification This procedure, designated as a catalyzed reporter deposition (CARD) or tyramide signal amplification (TSA), takes advantage of horseradish peroxidase (HRP) from an HRP-labeled secondary antibody to catalyze in the presence of hydrogen peroxide the oxidation of the phenol moiety of labeled tyramine. On oxidation by HRP, activated tyramine molecules rapidly bind covalently to electron-rich amino acids of proteins immediately surrounding the site of the immunoreaction. This allows an increase in the detection of an antigenic site up to 100-fold compared with the conventional indirect method with no loss in resolution. 

During this step, biotin sites on bound tyramine act as additional binding sites for antibiotin enzyme. An additional round of signal amplification can be achieved by using biotinyltyramide and streptavidin conjugated to alkaline phosphatase (Yang et al., 1999). 

Kerstens, H. M. J., Poddighe, P. J., and Hanselaar, A. G. J. M. 1995. A novel in situ hybridization signal amplification method based on the deposition of biotinylated tyramine. J. Histochem. Cytochem. 43 341-352. 

FIGURE 1.12 (A) Radicalization of tyramine amplification. (B) Tyramine-reporter deposition. (C) Comparison with the ABC method. From Merz H, Ottesen K, Meyer W, et al. Combination of antigen retrieval techniques and signal amplification of immunohistochemistry in situ hybridization and FISH techniques. In Shi S-R, Ciu I, Taylor CR, eds. Antigen Retrieval Techniques Immunohistochemistry and Molecular Morphology. Natick, MA Eaton 2000 228-229. 

---