A novel, automated technology for multiplex biomarker imaging and application to breast cancer

Area of Science:

• Oncology
• Biotechnology
• Pathology

Background:

• Multiplexed immunofluorescence (MIF) is crucial for validating multigene assays and understanding complex protein interactions in breast cancer.
• Current methods face challenges in simultaneously analyzing numerous biomarkers within a single tissue sample.

Purpose of the Study:

• To describe a novel technology for imaging an extended panel of biomarkers on a single formalin-fixed paraffin-embedded (FFPE) breast sample.
• To evaluate biomarker interactions at a single-cell level.
• To demonstrate proof-of-concept in breast tumors co-expressing hormone receptors, Her2/neu, and Ki-67.

Main Methods:

• Utilized a microfluidic flow cell for automated reagent exchange, including serial staining, imaging, and chemical deactivation.
• Developed a two-step antigen retrieval process to accommodate all epitopes simultaneously.
• Applied the imaging sequence to seven breast tumors and compared results with conventional immunohistochemistry (IHC).
• Performed single-cell correlation analysis using automated image processing.

Main Results:

• The novel platform successfully imaged an extended panel of biomarkers on FFPE breast tissue.
• Expression patterns in multiplexed images were consistent with conventional IHC.
• Automated processes reduced staining inconsistencies and positional shifts.
• Fluorescent dye cycling expanded the number of quantifiable biomarkers on a single section.

Conclusions:

• A novel platform for evaluating biomarker co-localization in breast cancer has been developed.
• The automated MIF technology enhances accuracy and efficiency in biomarker analysis.
• This approach significantly expands the capability to visualize and quantify multiple biomarkers on a single tissue section.