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Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction.

Publication ,  Journal Article
Kennedy-Darling, J; Bhate, SS; Hickey, JW; Black, S; Barlow, GL; Vazquez, G; Venkataraaman, VG; Samusik, N; Goltsev, Y; Schürch, CM; Nolan, GP
Published in: European journal of immunology
May 2021

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.

Duke Scholars

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Published In

European journal of immunology

DOI

EISSN

1521-4141

ISSN

0014-2980

Publication Date

May 2021

Volume

51

Issue

5

Start / End Page

1262 / 1277

Related Subject Headings

  • Single-Cell Analysis
  • Sensitivity and Specificity
  • Reproducibility of Results
  • Organ Specificity
  • Oligonucleotides
  • Molecular Imaging
  • Lymphoid Tissue
  • In Situ Hybridization
  • Immunology
  • Humans
 

Citation

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Kennedy-Darling, J., Bhate, S. S., Hickey, J. W., Black, S., Barlow, G. L., Vazquez, G., … Nolan, G. P. (2021). Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction. European Journal of Immunology, 51(5), 1262–1277. https://doi.org/10.1002/eji.202048891
Kennedy-Darling, Julia, Salil S. Bhate, John W. Hickey, Sarah Black, Graham L. Barlow, Gustavo Vazquez, Vishal G. Venkataraaman, et al. “Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction.European Journal of Immunology 51, no. 5 (May 2021): 1262–77. https://doi.org/10.1002/eji.202048891.
Kennedy-Darling J, Bhate SS, Hickey JW, Black S, Barlow GL, Vazquez G, et al. Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction. European journal of immunology. 2021 May;51(5):1262–77.
Kennedy-Darling, Julia, et al. “Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction.European Journal of Immunology, vol. 51, no. 5, May 2021, pp. 1262–77. Epmc, doi:10.1002/eji.202048891.
Kennedy-Darling J, Bhate SS, Hickey JW, Black S, Barlow GL, Vazquez G, Venkataraaman VG, Samusik N, Goltsev Y, Schürch CM, Nolan GP. Highly multiplexed tissue imaging using repeated oligonucleotide exchange reaction. European journal of immunology. 2021 May;51(5):1262–1277.
Journal cover image

Published In

European journal of immunology

DOI

EISSN

1521-4141

ISSN

0014-2980

Publication Date

May 2021

Volume

51

Issue

5

Start / End Page

1262 / 1277

Related Subject Headings

  • Single-Cell Analysis
  • Sensitivity and Specificity
  • Reproducibility of Results
  • Organ Specificity
  • Oligonucleotides
  • Molecular Imaging
  • Lymphoid Tissue
  • In Situ Hybridization
  • Immunology
  • Humans