iCLAP: an innovative method for integrable co-detection of low-abundance antigens with high-plex immunostaining

Area of Science:

• Biotechnology
• Molecular Biology
• Pathology

Background:

• Multiplexed protein imaging is crucial for spatial analysis of complex tissues.
• Detecting low-abundance proteins in formalin-fixed, paraffin-embedded (FFPE) tissues is challenging with current high-plex methods.
• Many important regulators like transcription factors and senescence markers are difficult to study in situ.

Purpose of the Study:

• To develop a sensitive and highly multiplexed protein detection method for FFPE tissues.
• To enable the study of low-abundance proteins in situ within FFPE specimens.
• To expand the capabilities of spatial proteomic analysis in FFPE tissues.

Main Methods:

• Introduced integrable Co-detection of Low-Abundant Proteins (iCLAP).
• iCLAP combines iterative signal amplification with efficient fluorophore inactivation.
• Enabled repeated staining on the same FFPE tissue section for over 40 markers.

Main Results:

• Demonstrated sensitive and highly multiplexed protein detection in FFPE tissues.
• Successfully profiled more than 40 markers on a single FFPE tissue section.
• Revealed spatially distinct senescence-associated protein patterns in human pancreatic tissues.

Conclusions:

• iCLAP significantly enhances the sensitivity of protein detection in FFPE tissues.
• The method facilitates high-dimensional spatial proteomic studies of complex biological processes.
• iCLAP expands the analytical utility of FFPE specimens for in situ protein analysis.