Sequential immunohistochemistry: a promising new tool for the pathology laboratory

Area of Science:

• Histopathology
• Immunohistochemistry
• Oncology

Background:

• Conventional immunohistochemistry (IHC) has limitations for multi-protein analysis in single tissue sections.
• Sequential IHC (S-IHC) offers a novel solution for studying numerous proteins within one section with minimal optimization.

Purpose of the Study:

• To introduce and validate sequential immunohistochemistry (S-IHC) as an improved method for multi-protein analysis.
• To assess the utility of S-IHC in characterizing lymphomas.

Main Methods:

• Tissue sections were stained sequentially with multiple antibodies, with intermediate scanning and elution of reagents.
• Digital images were overlaid to enable simultaneous assessment of multiple protein expressions.
• S-IHC was applied to nine nodular lymphocyte-predominant Hodgkin lymphomas (NLPHLs) and ten T-cell-rich/histiocyte-rich diffuse large B-cell lymphomas (T/HRBCLs).

Main Results:

• Expression of cyclin D1, CD20, and CD68 was analyzed in lymphoma subtypes.
• Cyclin D1 was detected in 44% of NLPHLs and 60% of T/HRBCLs.
• S-IHC identified discrepancies in 42% of cases compared to single IHC, underscoring challenges in distinguishing tumor cells from histiocytes.

Conclusions:

• S-IHC is a valuable technique for research and diagnostics, enabling comprehensive protein expression analysis in single tissue sections.
• The method preserves tissue architecture, facilitating detailed phenotypic characterization of individual cells.
• S-IHC demonstrates significant added value in complex cases like lymphomas where cell morphology can be ambiguous.