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Karkmann et al. introduced in 1999 an other method to enhance the
sensitivity of the intracellular staining protocol by using intracellular
tyramine-based signal amplification. They introduced it as the intracellular
tyramine-based signal of intracellular antigens (ITCA).
The saponin protocol
has been extended with the use of biotin ( instead of fluorochrome) labeled
antibodies to detect the molecules in the cell. After knocking out the
endogenous peroxidases and biotin residues the tyramine reaction makes it
possible to enhance the signal with a factor 15. Where the regular protocol
needs several thousands target molecules this protocol already is able to show
300 - 400 molecules per cell.
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ITCA
