Individual or pooled single-cell suspensions from thymus or spleen were obtained as previously described.26,30,31 (link) Axillary and inguinal lymph nodes were collected and mechanically dissected before counting the cells and staining them with flow cytometry antibodies. Cell counts were performed by Z2 particle counter (Beckman Coulter, Pasadena, CA), Spark 10M (Tecan, Zürich, Switzerland), or hemocytometer. CD45− cells were enriched by magnetic bead separation using LS columns and CD45 beads (Miltenyi Biotech,). Peripheral blood was collected into EDTA capillary pipettes (Drummond Scientific, Broomall, PA). Peripheral blood counts were performed on Element Ht5 automatic counter (Heska, Loveland, CO).
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Isolation and Characterization of Immune Cells
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Antibodies
Axillary
Blood
Capillary
Cells
Edta
Flow cytometry
Inguinal
Lymph nodes
Peripheral blood counts
Spleen
Thymus
Corresponding Organization :
Other organizations : Fred Hutch Cancer Center, University of Pisa, University of Washington, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, University of Siena
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Variable analysis
independent variables
- Tissue type (thymus, spleen, axillary/inguinal lymph nodes, peripheral blood)
- Cell counts
- Cell staining with flow cytometry antibodies
- Previously described methods for obtaining individual or pooled single-cell suspensions
- Magnetic bead separation using LS columns and CD45 beads for CD45- cell enrichment
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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