Profiling Immune Cell Populations in Inguinal Lymph Nodes

Draining inguinal lymph nodes from immunized mice were collected and treated as previously described (47 (link), 48 (link)). Briefly, the tissues were treated with collagenase type IV (Worthington) at a concentration of 1 mg/mL for 20 min at a temperature of 37 °C, and then passed through a 100-μm strainer to obtain a single-cell suspension. The resulting single-cell samples were stained with a range of markers including Zombie UV (BUV496, BioLegend 423107), anti-Ly6C (BV780, BioLegend 128041), anti-Ly6G (APC-Cy7, BioLegend 127624), anti-CD19 (BUV395, BD 563557), anti-CD3 (BB700, BD742175), anti-MHCII (AF700, eBioscience 56-5321-82), anti-CD11b (BV650, BioLegend 101239), anti-CD11c (BV421, BioLegend 117330), anti-CD86 (A647, BioLegend 105020), anti-Siglec-F (PE-CF594, BD 562757), anti-CD45 (BV610, BioLegend 103140), anti-CD169 (PE-Cy7, BioLegend 142412), anti-PDCA-1 (BUV563, BD 749275), anti-CD8a (BUV805, BD 612898), anti-CD103 (PE, eBioscience 12-1031-82), anti-NK1.1 (BV510, BioLegend 108738), and anti-F4/80 (BUV737, BD 749283). The cells were analyzed using the BD FACSymphony analyzer located at the Stanford Shared Fluorescence-activated cell sorting (FACS) Facility.

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Amaya L., Grigoryan L., Li Z., Lee A., Wender P.A., Pulendran B, & Chang H.Y. (2023). Circular RNA vaccine induces potent T cell responses. Proceedings of the National Academy of Sciences of the United States of America, 120(20), e2302191120.

Publication 2023

collagenase type IV BUV496 BV780 APC-Cy7 BUV395 BB700 AF700 BV650 BV421 PE-CF594 BV610 PE-Cy7 BUV563 BUV805 BV510 BUV737 BD FACSymphony analyzer

Anti cd3 Cd103 Cd11b Cells Inguinal Lymph nodes Mice Pdca 1 Siglec Tissues Type iv collagenase

Corresponding Organization :

Other organizations : Stanford University

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Variable analysis

independent variables

Collagenase type IV (Worthington) concentration (1 mg/mL)
Incubation time (20 min)
Incubation temperature (37 °C)

dependent variables

Populations of immune cells (e.g., Ly6C, Ly6G, CD19, CD3, MHCII, CD11b, CD11c, CD86, Siglec-F, CD45, CD169, PDCA-1, CD8a, CD103, NK1.1, F4/80)

control variables

Tissue source (draining inguinal lymph nodes from immunized mice)
Cell preparation method (passing through 100-μm strainer to obtain single-cell suspension)

controls

No positive or negative controls were explicitly mentioned in the provided information.

Annotations

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