Journal: Cell Death and Differentiation
Article Title: A defined metabolic state in pre B cells governs B-cell development and is counterbalanced by Swiprosin-2/EFhd1
doi: 10.1038/cdd.2017.52
Figure Lengend Snippet: Contribution of EFhd1tg precursors to B-cell development in mixed bone marrow chimeras. CD45.1 BL6 (Ly5.1) recipient mice were lethally irradiated with 11 Gy and reconstituted with 1 × 106 WT CD45.1 (Ly5.1) and 1 × 106 CD45.2 (Ly5.2) EFhd1tg or wild-type liitermate BM cells. After 6 weeks repopulation, the contribution of CD45.1 and CD45.2 to B-cell populations in the BM and spleen was analyzed by flow cytometry for pro B (CD19+, cKit+), pre B (CD19+, CD25+), immature (CD19+, IgM+, IgD−), mature (CD19+, IgM+, IgD+), marginal zone (CD19+, CD21high, CD23−) and follicular (CD19+, CD21low, CD23+) B cells. (a) Representative analysis of WT/EFhd1tg chimeras. (b) Summary of WT/EFhd1tg chimeras. (c) Summary of WT/WT chimeras. Data are represented as mean+S.E.M. of 7–11 mice from four experiments. Statistics was done by Mann–Whitney-U test
Article Snippet: The following antibodies were used for flow cytometry and cell sorting: unconjugated anti-CD16/32 (FC Block; 93; eBioscience), Alexa fluor 647 (AF647)-conjugated CD19 (eBio1D3; eBioscience), CD2 phycoerythrin (PE)-conjugated anti-CD19 (eBio1D3; eBioscience), Brilliant Violet 421 (BV)-conjugated anti-CD19 (BioLegend), fluorescein (FITC)-conjugated anti-CD21/35 (CR2/CR1; eBio8D9; eBioscience), FITC-conjugated anti-CD23 (eBioscience), PE-conjugated anti-CD23 (FcεRII; B3B4; eBioscience), APC-conjugated anti-CD25 (PC61; BioLegend), FITC-conjugated anti-CD43 (Ly-48, S7; BD), peridinin-chlorophyll protein-cyanine5.5 (PerCP)-conjugated anti-CD45R (B220; eBioscience), PE-conjugated anti-CD117 (c-kit; ACK45; BD), APC-conjugated anti-CD117 (c-kit, eBioscience), BV 421-conjugated anti-CD117 (c-kit, eBioscience), PerCP-Cy5.5-conjugated antiCD45.2 (eBioscience), Pacific Blue (PB)-conjugated anti-CD45.1 (BioLegend), Biotin-conjugated anti-IgD (Southern Biotech, distributed by Biozol, Eching, Germany), FITC-conjugated anti-IgM (Southern Biotech, distributed by Biozol, Eching, Germany), and Cyanine (Cy)5-conjugated anti-IgM (Southern Biotech); to detect biotinylated antibodies, PerCP-conjugated streptavidin (BD) were used.
Techniques: Irradiation, Flow Cytometry, MANN-WHITNEY