A 34-Marker Panel for Imaging Mass Cytometric Analysis of Human Snap-Frozen Tissue

Imaging mass cytometry (IMC) is able to quantify the expression of dozens of markers at sub-cellular resolution on a single tissue section by combining a novel laser ablation system with mass cytometry. As such, it allows us to gain spatial information and antigen quantification , and can be applied...

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Bibliographic Details
Published in:Frontiers in immunology Vol. 11; p. 1466
Main Authors: Guo, Nannan, van Unen, Vincent, Ijsselsteijn, Marieke E., Ouboter, Laura F., van der Meulen, Andrea E., Chuva de Sousa Lopes, Susana M., de Miranda, Noel F. C. C., Koning, Frits, Li, Na
Format: Journal Article
Language:English
Published: Switzerland Frontiers Media S.A 16.07.2020
Subjects:
Actins - metabolism
Adult
Antibodies - immunology
Antibodies - metabolism
Biomarkers - metabolism
Cadherins - metabolism
Female
Frozen Sections
human intestine
Humans
Image Cytometry - methods
imaging mass cytometry
IMC
Immunity, Innate
Immunology
Intestinal Mucosa - metabolism
Intestinal Mucosa - pathology
Lymphocytes - immunology
mass cytometry
Paraffin Embedding
snap-frozen tissue sections
T-Lymphocytes - immunology
Vimentin - metabolism
human intestine
imaging mass cytometry
snap-frozen tissue sections
mass cytometry
IMC
ISSN:1664-3224, 1664-3224
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Summary:Imaging mass cytometry (IMC) is able to quantify the expression of dozens of markers at sub-cellular resolution on a single tissue section by combining a novel laser ablation system with mass cytometry. As such, it allows us to gain spatial information and antigen quantification , and can be applied to both snap-frozen and formalin-fixed, paraffin-embedded (FFPE) tissue sections. Herein, we have developed and optimized the immunodetection conditions for a 34-antibody panel for use on human snap-frozen tissue sections. For this, we tested the performance of 80 antibodies. Moreover, we compared tissue drying times, fixation procedures and antibody incubation conditions. We observed that variations in the drying times of tissue sections had little impact on the quality of the images. Fixation with methanol for 5 min at -20°C or 1% paraformaldehyde (PFA) for 5 min at room temperature followed by methanol for 5 min at -20°C were superior to fixation with acetone or PFA only. Finally, we observed that antibody incubation overnight at 4°C yielded more consistent results as compared to staining at room temperature for 5 h. Finally, we used the optimized method for staining of human fetal and adult intestinal tissue samples. We present the tissue architecture and spatial distribution of the stromal cells and immune cells in these samples visualizing blood vessels, the epithelium and lamina propria based on the expression of α-smooth muscle actin (α-SMA), E-Cadherin and Vimentin, while simultaneously revealing the colocalization of T cells, innate lymphoid cells (ILCs), and various myeloid cell subsets in the lamina propria of the human fetal intestine. We expect that this work can aid the scientific community who wish to improve IMC data quality.
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Edited by: Christoph Mueller, University of Bern, Switzerland
These authors have contributed equally to this work
This article was submitted to Mucosal Immunity, a section of the journal Frontiers in Immunology
Reviewed by: Ruben Casanova, University of Zurich, Switzerland; Diane Bimczok, Montana State University, United States
ISSN:1664-3224
1664-3224
DOI:10.3389/fimmu.2020.01466