Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

Luke T Quigley; Lokman Pang; Elnaz Tavancheh; Matthias Ernst; Andreas Behren; Jennifer Huynh; Jessica Da Gama Duarte

doi:10.1016/j.xpro.2022.101961

Protocol for investigating tertiary lymphoid structures in human and murine fixed tissue sections using Opal™-TSA multiplex immunohistochemistry

STAR Protoc. 2023 Mar 17;4(1):101961. doi: 10.1016/j.xpro.2022.101961. Epub 2023 Jan 10.

Authors

Luke T Quigley¹, Lokman Pang¹, Elnaz Tavancheh¹, Matthias Ernst¹, Andreas Behren², Jennifer Huynh¹, Jessica Da Gama Duarte³

Affiliations

¹ Olivia Newton-John Cancer Research Institute, and School of Cancer Medicine, La Trobe University, Heidelberg, VIC 3084, Australia.
² Olivia Newton-John Cancer Research Institute, and School of Cancer Medicine, La Trobe University, Heidelberg, VIC 3084, Australia; Department of Medicine, University of Melbourne, Parkville, VIC 3010, Australia.
³ Olivia Newton-John Cancer Research Institute, and School of Cancer Medicine, La Trobe University, Heidelberg, VIC 3084, Australia. Electronic address: jessica.duarte@onjcri.org.au.

Abstract

Tertiary lymphoid structures (TLSs) are ectopic lymphoid aggregates that can develop within or adjacent to tumors, but protocols that can accurately identify and characterize TLSs are lacking. Here, we present a protocol for the in situ interrogation and characterization of TLSs in human and murine tissue sections using Opal™-tyramide signal amplification multiplex immunohistochemistry. This protocol enables simultaneous detection of up to 7 markers (6 antigens and a DAPI counterstain). We also describe a grading system to identify immature and mature TLSs.

Keywords: Cancer; Immunology; Microscopy.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Humans
Immunohistochemistry
Mice
Neoplasms* / pathology
Tertiary Lymphoid Structures* / pathology