Journal: bioRxiv

Article Title: Obesity alters mobility and adult neurogenesis, but not hippocampal dependent learning in ob/ob mice

doi: 10.1101/537720

Figure Lengend Snippet: Adult neurogenesis, proliferation and apoptosis were analyzed using immunofluorescence staining within the dentate gyrus (DG) of the hippocampus. Cell numbers of wild type (wt) animals were set to 100 %. A The number of proliferating cells was visualized using phosphohistone H3 (PH3). Leptin-deficient (ob/ob) mice showed a reduced amount of PH3-positive proliferating cells. B As a maker for immature neuronal cells, doublecortin (DCX) was used. Ob/ob mice show a reduced number of DCX positive cells in the DG compared to wild type controls. C As an apoptosis marker caspase3 (Casp3) was used. The number of Casp3 positive cells did not differ between the two groups of mice. Fluorescence images were taken using a Olympus BX63 microscope with 25,2x magnification (40x objective, 0,63x camera) in the DG of wt mice for D PH3 positive, E DCX positive and F Casp3 positive cells.

Article Snippet: After washing with PBS, the solution containing the primary rabbit anti-active Caspase 3 antibody (Merck Millipore, Germany) was applied 1:100 in blocking serum (3 % NGS + 0.1 % Triton X-100 in PBS) and incubated at 4°C over night.

Techniques: Immunofluorescence, Staining, Marker, Fluorescence, Microscopy

Journal: Journal of Immunology (Baltimore, Md. : 1950)

Article Title: Mechanism for Pre-B Cell Loss in V H -Mutant Rabbits

doi: 10.4049/jimmunol.1101778

Figure Lengend Snippet: Flow cytometric analysis of ali/ali and wt B-lineage cell survival 2–6 d after transfer of pro-B cells to OP9 stroma in the absence of IL-7. A, Relative number of B-lineage (CD79a+) cells normalized to ali/ali cultures using bead-based counting and gating strategy in Fig. 2A. Error bars show SD from three independent experiments. B, Live/Dead staining of total lymphocytes. C, Cross-reactivity of anti–caspase-3 Ab with a UV-irradiated rabbit B cell line (55D1). Left panel, UV-treated 55D1 B cells immediately after irradiation (t0) (gray filled) or 3 h postirradiation stained with anti–caspase-3 (black dashed line) or isotype control for 3 h time point (solid black line) (identical result was observed for isotype control staining at t0); number is percentage of caspase-3+ cells at 3 h. Right panel, CD79a expression in caspase-3+ (dashed line) and caspase-3neg (solid line) cells 3 h postirradiation. D, Frequency of active caspase-3+ B-lineage cells (CD79a+). Data for B and D are representative of three independent experiments.

Article Snippet: Abs and flow cytometry Abs used: mouse anti-human CD79a-PE (catalog number 555935) or APC (catalog number 551134), rabbit anti-human caspase-3–FITC (catalog number 559341), mouse anti-human spleen tyrosine kinase (Syk)-P-Y348 (I1s0-722), mouse anti-rabbit–μ (367.2-bio), and rat anti-mouse pre-BCR (SL-156-bio) from BD Pharmingen (San Diego, CA); rat anti-mouse–μ-PE and PE-Cy7 (II/41) from eBioscience (San Diego, CA); goat F(ab′) 2 anti-human IgH+L, rabbit F(ab)-FITC anti-goat IgG H+L, and allophycocyanin-conjugated streptavidin from Jackson ImmunoResearch Laboratories (West Grove, PA); and biotinylated rabbit anti–a2-allotype and FITC-rabbit anti–a2-allotype IgG (generated by K.L.K.’s laboratory).

Techniques: Staining, Irradiation, Expressing