Murine Splenic Cell Isolation

Spleens were aseptically harvested from euthanized mice. forced through 70-μm cell strainers (BD) with the plunger from a 3 mL syringe (BD). Cells were washed twice in cold RPMI or PBS followed by 5 minutes centrifugation at 1800 rpm. Cells were finally resuspended in supplemented RPMI medium containing 10% fetal calf serum (FCS) as previously described (34 (link)). Cells were counted using an automatic Nucleocounter (Chemotec) and cell suspensions were adjusted to 2×10⁵ cells/well for ELISA and 1–2×10⁶ cells/well for flow cytometry. and serum collection.

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Woodworth J.S., Contreras V., Christensen D., Naninck T., Kahlaoui N., Gallouët A.S., Langlois S., Burban E., Joly C., Gros W., Dereuddre-Bosquet N., Morin J., Olsen M.L., Rosenkrands I., Stein A.K., Wood G.K., Follmann F., Lindenstrøm T., LeGrand R., Pedersen G.K, & Mortensen R. (2023). A novel adjuvant formulation induces robust Th1/Th17 memory and mucosal recall responses in Non-Human Primates. bioRxiv.

Publication Preprint 2023

70-μm cell strainers 3 mL syringe

Cells Centrifugation Cold Elisa Fetal calf serum Flow cytometry Mice Serum Syringe

Corresponding Organization : Statens Serum Institut

Other organizations : Université Paris-Saclay, CEA Paris-Saclay, Commissariat à l'Énergie Atomique et aux Énergies Alternatives, Inserm, Infectious Disease Models and Innovative Therapies, CEA Paris-Saclay - Etablissement de Fontenay-aux-roses

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Variable analysis

independent variables

Euthanasia of mice
Aseptic harvesting of spleens
Passing spleens through 70-μm cell strainers with a 3 mL syringe plunger
Washing cells twice in cold RPMI or PBS
Centrifugation at 1800 rpm for 5 minutes
Resuspension of cells in supplemented RPMI medium containing 10% fetal calf serum (FCS)

dependent variables

Cell count (using an automatic Nucleocounter)
Cell suspensions adjusted to 2×10^5 cells/well for ELISA and 1–2×10^6 cells/well for flow cytometry and serum collection

control variables

Aseptic technique
Use of cold RPMI or PBS for washing
Centrifugation conditions (1800 rpm for 5 minutes)
Supplemented RPMI medium containing 10% FCS

controls

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

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