1. Carboxypeptidase A5 identifies a novel mast cell lineage in the zebrafish providing new insight into mast cell fate determination
by J. Tristan Dobson, Jake Seibert, Evelyn M. Teh, Sahar Da'as, Robert B. Fraser, Barry H. Paw, Tong-Jun Lin, and Jason N. Berman
Blood
Volume 112(7):
October 1, 2008
©2008 by American Society of Hematology

2. Zebrafish MCs structurally and functionally resemble their mammalian counterparts.
Zebrafish MCs structurally and functionally resemble their mammalian counterparts. H&E staining. (A) Intestine. (B) Gill. Periodic acid-Schiff (PAS). (C) Intestine. (D) Gill. Toluidine blue. (E) Intestine. (F) Gill. MCs in each panel indicated by ▶. Transmission electron microscope (Phillips 300; FEI, Hillsboro, OR) images of (G) a zebrafish intestinal MC (20 000× magnification) and (H) a mouse bone marrow–derived MC (26 000× magnification). Zebrafish MCs demonstrate a positive reaction to (I) a polyclonal antibody raised against human CD117 (C-KIT) antigen (Dako Cytomation, Carpinteria, CA) and (J) a monoclonal anti–human MC tryptase antibody (gills; Dako Cytomation). Biotinylated Universal Linker (Dako Cytomation) secondary antibody and 3,3′-diaminobenzidine for chromogenic detection (hematoxylin counterstain; 100× objective). RNA ISH using digoxigenin-labeled RNA antisense probe to zebrafish cpa5 demonstrates positive staining in (K) intestinal MCs and (L) pancreas; MCs indicated by ▶. Adult zebrafish injected intraperitoneally with 10 μg of compound 48/80 demonstrate (N) MC degranulation compared with (M) saline-injected controls. PAS staining, 100× objective; MCs in each panel indicated by ▶. (O) Increased plasma tryptase levels compared with saline-injected control fish. Presented as means plus SEM of 3 experiments with 4 to 6 fish per group; *P < .05 (t test). Cytospin of FACS-sorted, FITC-labeled, Fast Red–stained cpa5+ cells isolated from zebrafish embryos at 7 dpf demonstrate morphology consistent with MCs. (P) Toluidine blue. (Q) Wright-Giemsa. (R) Green channel (FITC). (S) Red channel (Fast Red; also Figure S2). (A-F, I-N) Images obtained at 100×/1.3 NA oil-immersion objective with a Nikon Eclipse E600, Nikon DXM 1200 camera, and ACT-1 software (all Nikon, Tokyo, Japan). Panels assembled using Adobe Photoshop CS3 Extended version 10.0 (Adobe Systems, San Jose, CA). (P-S) Images obtained at 100×/1.4 NA oil immersion objective with a Zeiss Axioplan 2 (Zeiss, Jena, Germany) and Axiocam HRc digital camera, Axiovision 4.6 with multi-channel fluorescence module (Diagnostic Instruments, Sterling Heights, MI).
J. Tristan Dobson et al. Blood 2008;112:
©2008 by American Society of Hematology

3. cpa5 identifies zebrafish MC progenitors.
cpa5 identifies zebrafish MC progenitors. (A) Double whole-mount ISH using a digoxigenin-labeled RNA antisense probe to zebrafish cpa5 (blue) and FITC-labeled RNA anti-sense probe (red) to pu.1, mpo, l-plastin, and lysozyme C (see Figure S3) demonstrate coexpression of cpa5 in a proportion of cells: (i) tail, (ii) head/yolk sac (5× objective). Evidence of coexpression is shown by colocalization observed in higher-magnification images of selected cells: (iii) brightfield, (iv) fluorescence (10× objective). No colocalization is observed for fms and cebp-α: (i) brightfield, (ii) fluorescence (8× objective). (B) gata-2 and pu.1 are both required for the development of early MCs as evidenced by the absence of cpa5 expression in gata-2 and pu.1 morphants, whereas gata-1 morphants paradoxically demonstrate increased numbers of cpa5+ cells. Fog-1 is dispensable for early MC development as evidenced by wild-type cpa5 expression in fog-1 morphants. Compound gata-1/pu.1 morphants demonstrate an absence of cpa5 expression, whereas compound gata-1/fog-1 morphants show a dramatic increase in numbers of cpa5+ cells. Lateral views, anterior left and dorsal at the top (28 hpf, 5× objective). Inset boxes demonstrate a higher-magnification view of the tail and the region around the intermediate cell mass. (C) Proposed model of transcription factor interactions required for early MC development. Established interactions are represented by —, and potential interactions by …. All images obtained with Leica application suite version 2.4.OR (Leica Microsystems, Heerbrugg, Switzerland); figure panels assembled using Adobe Photoshop CS3 Extended version 10.0 (Adobe Systems).
J. Tristan Dobson et al. Blood 2008;112:
©2008 by American Society of Hematology