Highly multiplexed immunofluorescence imaging of human tissues and tumors using t-CyCIF and conventional optical microscopes

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Cite this article as: eLife 2018;7:e31657 doi: 10.7554/eLife.31657

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Abstract

The architecture of normal and diseased tissues strongly influences the development and progression of disease as well as responsiveness and resistance to therapy. We describe a tissue-based cyclic immunofluorescence (t-CyCIF) method for highly multiplexed immuno-fluorescence imaging of formalin-fixed, paraffin-embedded (FFPE) specimens mounted on glass slides, the most widely used specimens for histopathological diagnosis of cancer and other diseases. t-CyCIF generates up to 60-plex images using an iterative process (a cycle) in which conventional low-plex fluorescence images are repeatedly collected from the same sample and then assembled into a high dimensional representation. t-CyCIF requires no specialized instruments or reagents and is compatible with super-resolution imaging; we demonstrate its application to quantifying signal transduction cascades, tumor antigens and immune markers in diverse tissues and tumors. The simplicity and adaptability of t-CyCIF makes it an effective method for pre-clinical and clinical research and a natural complement to single-cell genomics.

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Author details

Jia-Ren Lin

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Benjamin Izar

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Shu Wang

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Clarence Yapp

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Shaolin Mei

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Parin M Shah

Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, United States

Competing interests
No competing interests declared.
Sandro Santagata

Laboratory of Systems Pharmacology, Harvard Medical School, Boston, United States

Competing interests
No competing interests declared.
Peter K Sorger

Department of Systems Biology, Harvard Medical School, Boston, United States

For correspondence
peter_sorger@hms.harvard.edu

Competing interests
Peter K Sorger, PKS is a member of the Board of Directors of RareCyte Inc., which manufactures the slide scanner used in this study, and co-founder of Glencoe Software, which contributes to and supports open-source OME/OMERO image informatics software. Other authors have no competing financial interests to disclose..

"This ORCID iD identifies the author of this article:" 0000-0002-3364-1838

Funding

National Institutes of Health (P50GM107618)

Peter K Sorger

Dana-Farber/Harvard Cancer Center (GI SPORE Developmental Research Project Award)

Benjamin Izar

National Institutes of Health (U54HL127365)

Peter K Sorger

National Institutes of Health (R41-CA224503)

Peter K Sorger

Dana-Farber/Harvard Cancer Center (Claudia Adams Barr Program)

Benjamin Izar

National Institutes of Health (K08CA222663)

Benjamin Izar

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Human subjects: Formalin fixed and paraffin embedded (FFPE) tissues from were retrieved from the archives of the Brigham and Women's Hospital as part of discarded/excess tissue protocols or obtained from commercial vendors. The Institutional Review Board (IRB) of the Harvard Faculty of Medicine last reviewed the research described in this paper on 2/16/2018 (under IRB17-1688) and judged it to 'involve no more than minimal risk to the subjects' and thus eligible for a waiver of the requirement to obtain consent as set out in 45CFR46.116(d). Tumor tissue and FFPE specimens were collected from patients under IRB-approved protocols (DFCI 11-104) at Dana-Farber Cancer Institute/Brigham and Women's Hospital, Boston, Massachusetts. The consent waiver described above also covers these tissues and specimens.

Reviewing Editor

Arjun Raj, University of Pennsylvania, United States

Publication history

Received: September 1, 2017
Accepted: June 29, 2018
Accepted Manuscript published: July 11, 2018 (version 1)
Accepted Manuscript updated: July 12, 2018 (version 2)
Version of Record published: August 3, 2018 (version 3)

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.