A robust multiplex immunofluorescence and digital pathology workflow for the characterisation of the tumour immune microenvironment.

Molecular Oncology
Amélie Viratham PulsawatdiManuel Salto-Tellez

Abstract

Multiplex immunofluorescence is a powerful tool for the simultaneous detection of tissue-based biomarkers, revolutionising traditional immunohistochemistry. The Opal methodology allows up to eight biomarkers to be measured concomitantly without cross-reactivity, permitting identification of different cell populations within the tumour microenvironment. In this study, we aimed to validate a multiplex immunofluorescence workflow in two complementary multiplex panels and evaluate the tumour immune microenvironment in colorectal cancer (CRC) formalin-fixed paraffin-embedded tissue. We stained CRC and tonsil samples using Opal multiplex immunofluorescence on a Leica BOND RX immunostainer. We then acquired images on an Akoya Vectra Polaris and performed multispectral unmixing using inform. Antibody panels were validated on tissue microarray sections containing cores from six normal tissue types, using qupath for image analysis. Comparisons between chromogenic immunohistochemistry and multiplex immunofluorescence on consecutive sections from the same tissue microarray showed significant correlation (rs  > 0.9, P-value < 0.0001), validating both panels. We identified many factors that influenced the quality of the acquired fluorescent ...Continue Reading

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Citations

Dec 31, 2020·Clinical and Translational Medicine·Elisabeth StrackAndreas Weigert
Dec 31, 2020·Cancers·Fatima Abdullahi SidiManuel Salto-Tellez
Feb 19, 2021·Computational and Structural Biotechnology Journal·M P HumphriesM Salto-Tellez
Mar 16, 2021·Current Opinion in Oncology·Angélique Saint, Ellen Van Obberghen-Schilling
Mar 17, 2021·The Journal of Pathology·Lisa M McGinnisJames Ziai
Jun 6, 2021·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Leslie SolorzanoCarolina Wählby
Jun 27, 2021·NPJ Breast Cancer·Eileen E ParkesNiamh E Buckley
Jul 31, 2021·Cancer Cell·Emily F Davis-MarcisakLuciane T Kagohara
Sep 14, 2021·Molecular Oncology·Ulrik RingborgLaurence Zitvogel
Oct 31, 2021·Nature Protocols·Tariq Sami HaddadIris Nagtegaal
Nov 5, 2021·Modern Pathology : an Official Journal of the United States and Canadian Academy of Pathology, Inc·Xanthi StachteaDaniel B Longley

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Software Mentioned

StarDist
qupath
Akoya
Opal
Graphpad
dia
phenochart
prism
Nanostring
InSituPlex

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