Unlike some of the existing multiplex approaches such as nextgeneration sequencing, mPCR, mass spectrometry, etc. multiplex immunofluorescence -also referred to as fluorescent multiplex immunohistochemistry- provides an edge to understand the coexpression and spatial distribution of multiple targets without compromising the tissue integrity.
Tyramide Signal Amplification (TSA) is an immunostaining method designed for its high sensitivity, capable of detecting low expressed epitopes and generate precise and clear results. Tyramide is coupled to a fluorophore to generate a signal amplification. Via a reaction mediated by HRP (horseradish peroxidase) coupled with the secondary antibody (AbII), tyramide turns into its active conformation and covalently binds to tyrosine residues proximal to the target epitope1.
To detect the next antigen, an elution step is required in order to detach the primary and secondary antibody (AbI and AbII) complex from the epitope. The already coupled tyramide remains attached to the epitope proximal region due to the covalent bonding.
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