What are the Challenges Associated with Multiplexed Immunofluorescence?
- Antibody Cross-Reactivity: Non-specific binding of antibodies can lead to false-positive results, necessitating rigorous validation and optimization of antibody pairs. - Fluorescence Overlap: Spectral overlap between different fluorophores can complicate image analysis, requiring careful selection of fluorescence channels and the use of advanced imaging software. - Tissue Autofluorescence: Endogenous fluorescence from tissue components can interfere with signal detection, necessitating the use of autofluorescence quenching techniques.
