Novel In Situ Hybridization and Multiplex Immunofluorescence Technology Combined With Whole-slide Digital Image Analysis in Kidney Transplantation.
IF: 4.137
Cited by: 2


The elusive nature of assessing immunological processes in situ in organ transplantation is one of the major impediments to improve diagnostics and treatment. Here, we present a proof-of-concept study using multiplexed in situ hybridization (ISH) (RNAscope) to detect low-abundance cytokines in formalin-fixed paraffin-embedded (FFPE) human transplant kidney biopsies in combination with immunofluorescence (IF) for cell phenotyping. We show that a multiplex IF and ISH (mIFISH) assay is feasible to identify the cellular source of cytokines and chemokines (tumor necrosis factor-α, interferon-γ, and CXCL9) in FFPE transplant kidney biopsies and that quantification of the mRNA and protein signal is also possible at single-cell resolution in the context of tissue complexity. Furthermore, the mIFISH assay allows precise quantitative assessment of tubulitis, one of the key morphological correlates of alloimmune injury. Simultaneous in situ identification and quantification of multiple cellular phenotypes and mRNA expression of proinflammatory cytokines in FFPE tissues offer a novel insight into the biology of alloimmune injury in kidney transplantation and may contribute to improved diagnostic accuracy and patient care.


Gene Expression
acute cellular rejection
kidney transplantation
whole-slide digital image analysis

MeSH terms

Fluorescent Antibody Technique
In Situ Hybridization
Kidney Transplantation
Leukocyte Common Antigens
Molecular Imaging
Paraffin Embedding
Tissue Fixation


Junger, Henrik
Dobi, Dejan
Chen, Adeline
Lee, Linda
Vasquez, Joshua J
Tang, Qizhi
Laszik, Zoltan G

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