Automated quantitative high-throughput multiplex immunofluorescence pipeline to evaluate OXPHOS defects in formalin-fixed human prostate tissue
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Authors
Sachdeva, AshwinHart, Claire A
Carey, C. D.
Vincent, A. E.
Greaves, L. C.
Heer, R.
Oliveira, Pedro
Brown, Michael D
Clarke, Noel W
Turnbull, D. M.
Affiliation
Genito Urinary Cancer Research Group, Division of Cancer Sciences, Oglesby Cancer Research Building, University of Manchester, Manchester, M20 4GJ, UK.Issue Date
2022
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Advances in multiplex immunofluorescence (mIF) and digital image analysis has enabled simultaneous assessment of protein defects in electron transport chain components. However, current manual methodology is time consuming and labour intensive. Therefore, we developed an automated high-throughput mIF workflow for quantitative single-cell level assessment of formalin fixed paraffin embedded tissue (FFPE), leveraging tyramide signal amplification on a Ventana Ultra platform coupled with automated multispectral imaging on a Vectra 3 platform. Utilising this protocol, we assessed the mitochondrial oxidative phosphorylation (OXPHOS) protein alterations in a cohort of benign and malignant prostate samples. Mitochondrial OXPHOS plays a critical role in cell metabolism, and OXPHOS perturbation is implicated in carcinogenesis. Marked inter-patient, intra-patient and spatial cellular heterogeneity in OXPHOS protein abundance was observed. We noted frequent Complex IV loss in benign prostate tissue and Complex I loss in age matched prostate cancer tissues. Malignant regions within prostate cancer samples more frequently contained cells with low Complex I & IV and high mitochondrial mass in comparison to benign-adjacent regions. This methodology can now be applied more widely to study the frequency and distribution of OXPHOS alterations in formalin-fixed tissues, and their impact on long-term clinical outcomes.Citation
Sachdeva A, Hart CA, Carey CD, Vincent AE, Greaves LC, Heer R, et al. Automated quantitative high-throughput multiplex immunofluorescence pipeline to evaluate OXPHOS defects in formalin-fixed human prostate tissue. Vol. 12, Scientific Reports. Springer Science and Business Media LLC; 2022.Journal
Scientific ReportsDOI
10.1038/s41598-022-10588-zPubMed ID
35459777Additional Links
https://dx.doi.org/10.1038/s41598-022-10588-zType
ArticleLanguage
enae974a485f413a2113503eed53cd6c53
10.1038/s41598-022-10588-z
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