Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction
| Autor: | Gustavo Vazquez, Salil S. Bhate, Yury Goltsev, Garry P. Nolan, Sarah Black, Christian M. Schürch, Nikolay Samusik, Graham L. Barlow, Julia Kennedy-Darling, John W. Hickey, Vishal G. Venkataraaman |
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| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
In situ Cell type Lymphoid Tissue Immunology Oligonucleotides Cell Count Image processing Cell Communication Computational biology Biology Sensitivity and Specificity Stain Antibodies 03 medical and health sciences Spatial single‐cell biology 0302 clinical medicine Single-cell analysis DNA‐conjugated antibodies Humans Immunology and Allergy Basic Single‐cell analysis New technology In Situ Hybridization Tissue homeostasis CODEX Multiplexed tissue imaging Histocytochemistry Oligonucleotide Reproducibility of Results Molecular Imaging 3. Good health Staining 030104 developmental biology Organ Specificity Research Article|Basic Single-Cell Analysis Research Article 030215 immunology |
| Zdroj: | European Journal of Immunology |
| ISSN: | 1521-4141 0014-2980 |
| DOI: | 10.1002/eji.202048891 |
| Popis: | Multiparameter tissue imaging enables analysis of cell‐cell interactions in situ, the cellular basis for tissue structure, and novel cell types that are spatially restricted, giving clues to biological mechanisms behind tissue homeostasis and disease. Here, we streamlined and simplified the multiplexed imaging method CO‐Detection by indEXing (CODEX) by validating 58 unique oligonucleotide barcodes that can be conjugated to antibodies. We showed that barcoded antibodies retained their specificity for staining cognate targets in human tissue. Antibodies were visualized one at a time by adding a fluorescently labeled oligonucleotide complementary to oligonucleotide barcode, imaging, stripping, and repeating this cycle. With this we developed a panel of 46 antibodies that was used to stain five human lymphoid tissues: three tonsils, a spleen, and a LN. To analyze the data produced, an image processing and analysis pipeline was developed that enabled single‐cell analysis on the data, including unsupervised clustering, that revealed 31 cell types across all tissues. We compared cell‐type compositions within and directly surrounding follicles from the different lymphoid organs and evaluated cell‐cell density correlations. This sequential oligonucleotide exchange technique enables a facile imaging of tissues that leverages pre‐existing imaging infrastructure to decrease the barriers to broad use of multiplexed imaging. We simplified the multiplexed imaging method CO‐Detection by indEXing (CODEX) by employing cyclic stripping and annealing of fluorescent oligonucleotides to DNA‐barcoded antibodies. We stained five human lymphoid tissues with a panel of 46 antibodies. This enabled quantitative spatial single‐cell biology analysis of differential cell‐type composition of immune follicles. |
| Databáze: | OpenAIRE |
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