Isolation and Staining of Lymphoid Cells

Secondary lymphoid organs (spleen and lymph nodes-axillary, brachial, inguinal, cervical, mesenteric, para-aortic) were isolated from mice and were pressed through a 70-μm nylon screen to generate a single-cell suspension. Single-cell suspensions were stained for 1 h at room temperature with allophycocyanin- or phycoerythrin-conjugated tetramers. Samples underwent enrichment for tetramer-binding cells as previously described (57 (link)). Cre_61–71:I-A^b APC and PE tetramers were a generous gift from James Moon.

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Martinez R.J., Breed E.R., Worota Y., Ashby K.M., Vobořil M., Mathes T., Salgado O.C., O’Connor C.H., Kotenko S.V, & Hogquist K.A. (2023). Type III interferon drives thymic B cell activation and regulatory T cell generation. Proceedings of the National Academy of Sciences of the United States of America, 120(9), e2220120120.

Publication 2023

Allophycocyanin Aortic Axillary Cells Cervical Inguinal Lymph nodes Mesenteric Mice Nylon Para Phycoerythrin Secondary lymphoid organs Spleen Tetramers

Corresponding Organization :

Other organizations : University of Minnesota Medical Center, Rutgers, The State University of New Jersey

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Variable analysis

independent variables

Isolation of secondary lymphoid organs (spleen and lymph nodes-axillary, brachial, inguinal, cervical, mesenteric, para-aortic) from mice

dependent variables

Generation of single-cell suspensions from the isolated secondary lymphoid organs
Staining of single-cell suspensions with allophycocyanin- or phycoerythrin-conjugated tetramers
Enrichment for tetramer-binding cells

control variables

Nylon screen used to generate single-cell suspensions
Room temperature used for staining
Staining duration of 1 hour

positive controls

Cre61–71:I-Ab APC and PE tetramers provided by James Moon

negative controls

Not explicitly mentioned

Annotations

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