Light sheet fluorescence microscopy for in situ cell interaction analysis in mouse lymph nodes.

Abe, Jun; Ozga, Aleksandra Joanna; Swoger, Jim; Sharpe, James; Ripoll, Jorge; Stein, Jens Volker (2016). Light sheet fluorescence microscopy for in situ cell interaction analysis in mouse lymph nodes. Journal of immunological methods, 431, pp. 1-10. Elsevier 10.1016/j.jim.2016.01.015

[img] Text
1-s2.0-S0022175916300151-main.pdf - Published Version
Restricted to registered users only
Available under License Publisher holds Copyright.

Download (1MB) | Request a copy

Reactive lymph nodes (LNs) are sites where pMHC-loaded dendritic cells (DCs) interact with rare cognate T cells, leading to their clonal expansion. While DC interactions with T cell subsets critically shape the ensuing immune response, surprisingly little is known on their spatial orchestration at physiologically T cell low precursor frequencies. Light sheet fluorescence microscopy and one of its implementations, selective plane illumination microscopy (SPIM), is a powerful method to obtain precise spatial information of entire organs of 0.5-10mm diameter, the size range of murine LNs. Yet, its usefulness for immunological research has thus far not been comprehensively explored. Here, we have tested and defined protocols that preserve fluorescent protein function during lymphoid tissue clearing required for SPIM. Reconstructions of SPIM-generated 3D data sets revealed that calibrated numbers of adoptively transferred T cells and DCs are successfully detected at a single cell level within optically cleared murine LNs. Finally, we define parameters to quantify specific interactions between antigen-specific T cells and pMHC-bearing DCs in murine LNs. In sum, our studies describe the successful application of light sheet fluorescence microscopy to immunologically relevant tissues.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Pre-clinic Human Medicine > Theodor Kocher Institute

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Abe, Jun; Ozga, Aleksandra Joanna and Stein, Jens Volker

Subjects:

600 Technology > 610 Medicine & health

ISSN:

0022-1759

Publisher:

Elsevier

Language:

English

Submitter:

Ursula Zingg-Zünd

Date Deposited:

06 Jul 2016 10:56

Last Modified:

06 Jul 2016 10:56

Publisher DOI:

10.1016/j.jim.2016.01.015

PubMed ID:

26844990

Uncontrolled Keywords:

Light sheet fluorescence microscopy; Lymph nodes; T cell–DC interactions; Tissue clearing

BORIS DOI:

10.7892/boris.83980

URI:

https://boris.unibe.ch/id/eprint/83980

Actions (login required)

Edit item Edit item
Provide Feedback