5-FITC tyramide is a green fluorescent dye composed of FITC and tyramine, with excitation at 488 nm and emission at 525 nm. It serves as a fluorescent substrate for horseradish peroxidase (HRP)-catalyzed deposition, useful in tyramide signal amplification (TSA). 5-FITC tyramide is applicable in multiplex immunohistochemistry (mIHC) experiments.

5-FITC tyramide detection involves several key steps: 1) Inject 4 μL of WGA-HRP tracer into the rat ganglion, then collect specific tissues for sectioning. 2) Air-dry the sections for 60 minutes. 3) Rehydrate the sections in TNT buffer [0.1 M Tris-HCl, 0.15 M NaCl, 0.05% Tween 20, pH 7.5], followed by quenching in freshly prepared NaBH4 [0.5 mg/mL] in TN buffer [0.1 M Tris-HCl, 0.15 M NaCl, pH 7.5] for 15 minutes. 4) Wash with TNT and permeabilize with TN buffer containing 0.1% Triton X-100 for 15 minutes. 5) Incubate in TNT buffer with 2% BSA (TNT-BSA) for 60 minutes, then wash with TNT and incubate with 5-FITC tyramide [diluted 1:50 in an amplification buffer] for 15 minutes. 6) Wash with TNT, incubate with primary antibody overnight at 4°C, then add donkey anti-rabbit IgG conjugated with Lissamine rhodamine (LRSC) or Cy3 [diluted 1:200 in TNT-BSA] and donkey anti-goat conjugated Cy5 [diluted 1:200 in TNT-BSA]. 7) Finally, perform fluorescence microscopy detection. Note: The optimal concentration of 5-FITC tyramide working solution should be determined based on actual conditions.