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Validation of multiplex immunofluorescence panels using multispectral microscopy for immune-profiling of formalin-fixed and paraffin-embedded human tumor tissues.

Abstract：

:Immune-profiling is becoming an important tool to identify predictive markers for the response to immunotherapy. Our goal was to validate multiplex immunofluorescence (mIF) panels to apply to formalin-fixed and paraffin-embedded tissues using a set of immune marker antibodies, with the Opal™ 7 color Kit (PerkinElmer) in the same tissue section. We validated and we described two panels aiming to characterize the expression of PD-L1, PD-1, and subsets of tumor associated immune cells. Panel 1 included pancytokeratin (AE1/AE3), PD-L1, CD4, CD8, CD3, CD68, and DAPI, and Panel 2 included pancytokeratin, PD-1, CD45RO, granzyme B, CD57, FOXP3, and DAPI. After all primary antibodies were tested in positive and negative controls by immunohistochemistry and uniplex IF, panels were developed and simultaneous marker expressions were quantified using the Vectra 3.0™ multispectral microscopy and image analysis InForm™ 2.2.1 software (PerkinElmer).These two mIF panels demonstrated specific co-localization in different cells that can identify the expression of PD-L1 in malignant cells and macrophages, and different T-cell subpopulations. This mIF methodology can be an invaluable tool for tumor tissue immune-profiling to allow multiple targets in the same tissue section and we provide that is accurate and reproducible method when is performed carefully under pathologist supervision.

journal_name

Sci Rep

journal_title

Scientific reports

authors

Parra ER,Uraoka N,Jiang M,Cook P,Gibbons D,Forget MA,Bernatchez C,Haymaker C,Wistuba II,Rodriguez-Canales J

doi

10.1038/s41598-017-13942-8

subject

Has Abstract

pub_date

2017-10-17 00:00:00

pages

13380

issue

1

issn

2045-2322

pii

10.1038/s41598-017-13942-8

journal_volume

7

pub_type

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