Recent advances in multiplex immunohistochemistry techniques allow for quantitative, spatial identification of multiple immune parameters for enhanced diagnostic and prognostic insight. However, applying such techniques to murine fixed tissues, particularly sensitive epitopes, such as CD4, CD8α, and CD19, has been difficult. We compared different fixation protocols and Ag-retrieval techniques and validated the use of multiplex immunohistochemistry for detection of CD3+CD4+ and CD3+CD8+ T cell subsets in murine spleen and tumor. This allows for enumeration of these T cell subsets within immune environments, as well as the study of their spatial distribution.
The online version of this article contains supplemental material.
Abbreviations used in this article:
- dendritic cell
- double-distilled H2O
- neutral-buffered 10% formalin
- room temperature
- Received December 31, 2015.
- Accepted February 22, 2016.
- Copyright © 2016 by The American Association of Immunologists, Inc.
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