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Updated: May 16, 2026
Multiplex Immunohistochemistry Staining for Paraffin-embedded Lung Cancer Tissue
Published on: November 21, 2023
A simple and reliable immunohistochemical method for colocalization of 2 antigens in the same cells of
Jun Yan1, Vibeke S Catts, Anthony Chan
1UQ Centre for Clinical Research, University of Queensland, Brisbane, Qld, Australia.
Insights
A new microwave heating technique (MVT) allows efficient colocalization of multiple antigens in single cells on tissue sections. This cost-effective method enhances sensitivity and specificity for improved clinical diagnosis and research.
Area of Science:
- Biomedical research
- Histopathology
- Immunohistochemistry
Background:
- Current immunohistochemistry (IHC) methods struggle with efficient multi-antigen colocalization on single tissue sections.
- There is a need for cost-effective, sensitive, and specific techniques for simultaneous antigen detection.
Purpose of the Study:
- To develop a novel sequential staining technique for colocalizing two different antigens within the same cell in paraffin-embedded tissue sections.
- To improve upon existing methods by enhancing efficiency, cost-effectiveness, and diagnostic utility.
Main Methods:
- Developed a sequential double staining technique combining microwave heating technique (MVT) with standard IHC protocols.
- Utilized a digital management system for comparing results and colocalizing antigens in single cells.
- Antibodies could be from the same or different species.
Main Results:
- The MVT colocalization method demonstrated high sensitivity and specificity, comparable to conventional immunofluorescence and IHC.
- The technique allows for precise colocalization and comparison of cell morphology across two images.
- This method avoids uncertainties associated with overlapping antigens in single images.
Conclusions:
- The novel MVT sequential staining technique offers an inexpensive and convenient solution for multi-antigen colocalization in paraffin-embedded tissues.
- This method enhances diagnostic accuracy and research capabilities in histopathology.
- It provides a reliable alternative for visualizing multiple antigens in single cells.
Abstract:
Immunohistochemistry (IHC) lacks an efficient technique for colocalizing multiple antigens in the same cells of a single tissue section. The development of a methodology which combines the advantage of low cost, high sensitivity, and specificity would benefit clinical diagnosis and general research. On the basis of a newly published method of visualizing 2 antigens on a single paraffin-embedded tissue section, we have further developed a novel sequential technique for colocalizing 2 different antigens in a same cell in a paraffin-embedded tissue section. In this technique, we combined the microwave heating technique (MVT) with normal IHC methods to sequentially double stain a paraffin section; and colocalize 2 antigens in a single cell through result comparison stored in a digital management system. This MVT colocalization method has a higher degree of sensitivity and specificity comparable with conventional staining of both immunofluorescence and IHC systems. The primary advantage of this method is that it is inexpensive and convenient; the antibody(s) used in this method can be generated from the same or different species; it allows colocalization or comparison of different results of cell morphology for any single cell of the section on 2 images, avoids uncertainty when overlapping 2 antigens on a single image.
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